Reactivity and Mechanism of Manganese Enzymes - Advances in

10 Reactivity and Mechanism of Manganese Enzymes A Modeling Approach

Downloaded by UNIV OF MINNESOTA on May 13, 2013 | http://pubs.acs.org Publication Date: May 5, 1996 | doi: 10.1021/ba-1995-0246.ch010

Vincent L . Pecoraro, Andrew Gelasco, and Michael J. Baldwin The Willard H . Dow Laboratories, Department of Chemistry, The University of Michigan, Ann Arbor, MI 48109-1055

Manganese satisfies a wide variety of biological activities that range from Lewis acid-base reactions to redox catalysis. This chapter summarizes the proposed chemical mechanisms for a wide variety of manganese-based enzymes that fulfill both nonredox and redox roles. We follow this minireview of manganese enzymes with specific examples drawn from our laboratory showing how small molecule coordination complexes can be prepared that mimic structural, spectroscopic, and reactivity properties of two redox-based manganese enzymes: the manganese catalase and the oxygen evolving complex. We describe two relatively simple systems composed of manganese dimers that catalyze H O disproportionation using either low-valent (Mn(II) -> Mn(III) ) or high-valent(Mn(III) -> Mn(IV) ) cycles. From this work we illustrate how understanding the mechanism of well-defined model compounds may provide new mechanistic insight to more complex and ill-defined biological catalytic centers. 2

2

2

2

2

2

OVER THE PAST DECADE

inorganic chemists, biochemists, and biophysicists have gained an enhanced appreciation for the role of manganese in the biosphere. Although once a curiosity, manganese is now recognized to be an essential component in photosynthesis and is found to be an alternative to iron in a wide variety of redox enzymes. In addition, the metal is an excellent Lewis acid catalyst and often confers stability to protein structure. In this chapter we summarize the known chemistry of manganese in biological systems and in compounds that have been synthesized in our laboratory to mimic the reactivity of these fascinating 0065-2393/95/0246-0265/$ 11.24/0 © 1995 American Chemical Society

In Mechanistic Bioinorganic Chemistry; Thorp, H., et al.; Advances in Chemistry; American Chemical Society: Washington, DC, 1996.

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M E C H A N I S T I C BIOINORGANIC CHEMISTRY

p r o t e i n s as a n e x a m p l e o f t h e m o d e l i n g a p p r o a c h f o r u n d e r s t a n d i n g their mechanisms.

Biological

Chemistry of Manganese


Manganese plays an essential and versatile role i n the b i o c h e m i s t r y of m a n y m i c r o o r g a n i s m s , p l a n t s , a n d a n i m a l s . M a n g a n e s e is c o n s i d e r e d a n essential trace element i n humans w i t h total b o d y metal b e i n g ~ 2 0 m g ( i , 2). I n p l a n t s a n d o x y g e n - e v o l v i n g p h o t o s y n t h e t i c b a c t e r i a , m a n g a n e s e is a n e s s e n t i a l c o m p o n e n t o f t h e o x i d a t i v e e n d o f p h o t o s y n t h e s i s . N u m e r o u s b a c t e r i a l sources use manganese i n place o f i r o n i n reactions i n v o l v i n g o x y g e n d e t o x i f i c a t i o n . M a n g a n e s e is a n e x t r e m e l y v e r s a t i l e e l e m e n t i n b i o l o g y b e c a u s e t h i s e l e m e n t is r e l a t i v e l y a b u n d a n t , has m u l t i p l e a c c e s s i b l e r e d o x states, a n d , i n t h e + 2 o x i d a t i o n l e v e l , is s i m i l a r i n s i z e a n d p r o p e r t i e s t o M g ( I I ) . T h e r e f o r e , m a n g a n e s e m a y s e r v e as a L e w i s a c i d catalyst or be d i r e c t l y i n v o l v e d i n m u l t i e l e c t r o n redox c o n versions. In the following section we describe the v a r i e d functions of manganoenzymes and cofactors placing particular emphasis on the m e c h a n i s t i c aspects of this c h e m i s t r y . E n z y m e s that use manganese i n a p u r e l y structural role w i l l not be discussed. T h e s i m p l e s t u s e o f m a n g a n e s e is as p a r t o f a m e t a l c o f a c t o r s u c h as manganese(II) adenosine t r i p h o s p h a t e , [ M n ( I I ) A T P ] . T h e m e t a l c o m p l e x is u s u a l l y f o u n d as t h e b i d e n t a t e [ Μ η ( Ι Ι ) β , 7 - Α Τ Ρ ~ ] (3, 4) as i l l u s t r a t e d i n F i g u r e 1. E n z y m e s p e c i f i c i t y f o r M g ( I I ) , M n ( I I ) , o r C a ( I I ) A T P c o m p l e x e s is d e p e n d e n t o n a v a r i e t y o f f a c t o r s ; h o w e v e r , o n c e s e l e c t e d , each m e t a l a p p a r e n t l y functions i n an analogous m a n n e r . T h e m e t a l i o n serves t w o m a i n purposes. F i r s t , based o n the c o o r d i n a t i o n g e o m e t r y 2 -

2

OH

OH

Figure 1. β,y-bidentate Mn(II) adenosine triphosphate dianion. The Mn(II) coordination sphere includes four additional water molecules.


10.

267

Reactivity and Mechanism of Manganese Enzymes

PECORARO

of the complex, the phosphate oxygens and the b o u n d water molecules can f o r m h y d r o g e n bonds that stabilize a specific c o n f o r m a t i o n of the M ( I I ) A T P c o m p l e x . T h i s is i m p o r t a n t i n l e a d i n g t o p r o d u c t i v e b i n d i n g o f t h e M ( I I ) A T P " to a l l o w p h o s p h o r y l g r o u p t r a n s f e r . S e c o n d , t h e m e t a l p o l a r i z e s t h e O - P b o n d so as t o m o v e e l e c t r o n d e n s i t y a w a y f r o m t h e phosphorus atom. T h e generation of this δ makes the phosphorus a b e t t e r site for a t t a c k b y a n e n t e r i n g n u c l e o p h i l e s u c h as a n a l c o h o l , a m i n e , or p h e n o l . T h i s S 2 m e c h a n i s m leads to a t r i g o n a l b i p y r a m i d a l p h o s p h o r u s i n t e r m e d i a t e that c o l l a p s e s t o [ Μ η ( Π ) ( Η 0 ) ( β A D P ) ( 0 P O R ) ] " and subsequently forms i n solution the products O3POR - and [Mn(n)(H 0) (a,j8-ADP)]-. T h e reversibility of a kinase ( p h o s p h o r y l t r a n s f e r e n z y m e ) is c o r r e l a t e d w i t h t h e m e t a l A T P c o m p l e x t h a t is r e c o g n i z e d as a s u b s t r a t e (4). I r r e v e r s i b l e e n z y m e s s u c h as h e x o kinase (reaction: glucose + [ M ( I I ) A T P ] " glucose-6-phosphate + [ M ( I I ) A D P ) ] - ) u s e b i d e n t a t e [ Μ ( Ι Ι ) β , γ - Α Τ Ρ ] - as s u b s t r a t e i n t h e f o r w a r d reaction a n d monodentate [M(II)jS-ADP]" for the reverse r e a c t i o n . T h i s substrate preference strongly drives the reaction t o w a r d products b e c a u s e t h e e q u i l i b r i u m c o n c e n t r a t i o n o f [ Μ ( Ι Ι ) β , γ - Α Τ Ρ ] ~ is l a r g e , w h e r e a s t h e c o n c e n t r a t i o n o f [ M ( I I ) / 3 - A D P ) ] ~ is e x t r e m e l y s m a l l . I n c o n trast, r e v e r s i b l e e n z y m e s s u c h as c r e a t i n e k i n a s e ( r e a c t i o n : c r e a t i n e + [M(II)ATP] p h o s p h o c r e a t i n e + [ M ( I I ) A D P ) ] - ) use t r i d e n t a t e [Μ(ΙΙ)α,0,γ-ΑΤΡ] - i n the f o r w a r d reaction and [M(II)(a,j8-ADP)]- i n the r e v e r s e r e a c t i o n . I n t h i s case t h e c o n c e n t r a t i o n o f [ M ( I I ) ( a , / 3 - A D P ) ] ~ e x c e e d s that o f [α,β,γ-tridentate [ M ( I I ) A T P ] " . A n o t h e r e x a m p l e o f α , β , γ t r i d e n t a t e [ M ( I I ) A T P ] " is t h e 0 - s u b u n i t o f t h e c h l o r o p l a s t A T P s y n t h a s e , w h i c h a p p a r e n t l y r e c o g n i z e s t r i d e n t a t e M n ( I I ) A T P as s u b s t r a t e . 2 _

2

+

N

2

2


4

3

3

2

4

2

2

2

2

2

2

2

T h e i n t e r a c t i o n of manganese or m a g n e s i u m w i t h n u c l e i c acids p r o vides additional interesting examples of charge neutralization or polar i z a t i o n effects o f t h e t y p e d e s c r i b e d i n t h e p r e c e d i n g p a r a g r a p h . C e c h (5) has s h o w n t h a t m e t a l c o f a c t o r s are n e e d e d for t h e p r o p e r f u n c t i o n i n g o f ribozymes. I n a d d i t i o n to a p o s s i b l e s t r u c t u r a l r o l e , it is n o w e s t a b l i s h e d t h a t e i t h e r M n ( I I ) o r M g ( I I ) w i l l f u n c t i o n as e s s e n t i a l c o f a c t o r s f o r t h e c l e a v a g e o f e x o g e n o u s R N A o r D N A b y t h e Tetrahymena r i b o z y m e . A p p a r e n t l y , the M(II) c a t i o n b i n d s d i r e c t l y to the r i b o z y m e i n the active s i t e , s e r v i n g to s t a b i l i z e t h e n e g a t i v e c h a r g e o n t h e o x y g e n a t o m o f t h e 3 Ό - Ρ b o n d i n t h e t r a n s i t i o n state. T h u s , i t w a s c o n c l u d e d t h a t t h e r i b o z y m e s are m e t a l l o e n z y m e s that have m e c h a n i s t i c characteristics that a r e s i m i l a r to p r o t e i n e n z y m e s . A l t h o u g h M n ( I I ) m a y f o r m d i r e c t i n n e r s p h e r e c o m p l e x e s w i t h n u c l e o t i d e s , D N A , o r R N A , t h e r e a r e cases i n w h i c h c h a r g e , n o t L e w i s a c i d i t y , is t h e d o m i n a n t f a c t o r for c a t a l y s i s . C o w a n (6) s u g g e s t e d that i n s o m e cases, t h e o n l y a p p a r e n t f u n c t i o n o f t h e m e t a l is c h a r g e n e u t r a l i z a t i o n . I n t h i s case, t h e fact t h a t t h e t o p o isomerase I hydrolysis of D N A c o u l d be affected b y C o ( N H ) in place o f M g ( I I ) suggests that a n i n n e r s p h e r e c o m p l e x is n o t n e c e s s a r y f o r 3

6

3 +


268


catalysis. R a t h e r , the a p p r o p r i a t e functional groups can be o r i e n t e d i n c l o s e p r o x i m i t y o n c e t h e p h o s p h a t e b a c k b o n e is p a r t i a l l y n e u t r a l i z e d . A l t h o u g h the majority of proteins containing multinuclear centers c o m p o s e d partially or c o m p l e t e l y w i t h manganese are i n v o l v e d i n redox reactions, there are a significant n u m b e r of substrate b i n d i n g or h y d r o lytic activities that r e l y on m e t a l clusters that i n c l u d e manganese. Rat l i v e r arginase c a t a l y z e s t h e h y d r o l y s i s o f L - a r g i n i n e t o g i v e L - o r n i t h i n e a n d u r e a . M a n g a n e s e ( I I ) is t h e e s s e n t i a l c o f a c t o r f o r t h i s e n z y m e (7, S i , Si S , S S , S S , S c y c l e i n r i g o r o u s l y d a r k - a d a p t e d sites (25, 26). H o w e v e r a r e i n t e r p r e t a t i o n o f the p r o t o n release e x p e r i m e n t s w i t h N e u t r a l r e d has i m p l i c a t e d a s t r o n g l y p H - d e p e n d e n t n o n i n t e g e r s t o i c h i o m e t r y (27). I n y e t a n o t h e r e x p e r i m e n t J a h n s et a l . (28) f o u n d a p a t tern of 1:1:1:1. 0

2

2

3

3

4

0

T h e d a r k - s t a b l e r e s t i n g state is t h e Si o x i d a t i o n l e v e l . A s i n g l e flash o f l i g h t l e a d s t o t h e S state, w h i c h has b e e n i n t e n s i v e l y s t u d i e d b e c a u s e o f its ease o f access a n d E P R s p e c t r a l f e a t u r e s . T h e E P R s p e c t r u m o b t a i n e d at l i q u i d h e l i u m t e m p e r a t u r e s o f t h e S state s h o w s a 1 9 - 2 6 l i n e s i g n a l c e n t e r e d at g « 2. T h e b r e a d t h o f t h e s i g n a l , t h e m a g n i t u d e o f the nuclear hyperfine coupling, and the energy of the transition conc l u s i v e l y i d e n t i f i e s t h i s s i g n a l as a r i s i n g f r o m a m i x e d - v a l e n c e m a n g a n e s e c l u s t e r (29). T h e o b s e r v a t i o n o f t h i s s i g n a l w a s t h e first d e m o n s t r a t i o n t h a t at least s o m e o f t h e m a n g a n e s e f o r m a c l u s t e r i n S . T h e m u l t i l i n e E P R s i g n a l e x h i b i t s t h e s a m e f o u r - f l a s h p e r i o d i c i t y as o b s e r v e d f o r t h e o x y g e n y i e l d , b u t is d i s p l a c e d b y t w o o x i d i z i n g e q u i v a l e n t s w i t h i n t h e c y c l e (i.e., 0 p r o d u c t i o n f o l l o w s S - > S , w h e r e a s m u l t i l i n e i n t e n s i t y c y c l e s at S ) . T h i s o b s e r v a t i o n d i r e c t l y l i n k s t h e m a n g a n e s e i o n s t o t h e o x i d a t i v e c h e m i s t r y o f t h i s e n z y m e . R e c e n t X - r a y a b s o r p t i o n d a t a (30, 31) also i m p l i c a t e s m a n g a n e s e i n t h e r e d o x c y c l e . T h e p r e s e n t l y a c c e p t e d o x i d a t i o n states f o r t h e m a n g a n e s e i o n s i n e a c h o x i d a t i o n l e v e l , as w e l l as l i k e l y c a n d i d a t e s f o r m e t a l l i g a n d s a r e p r o v i d e d i n T a b l e I. 2

2

2

2

0

4

2

A n o t h e r E P R s i g n a l at g « 4.1 c a n b e o b s e r v e d u n d e r c e r t a i n c o n d i t i o n s (e.g., a d d i n g a m m o n i a t o t h e s a m p l e ) (32-36). T h i s s i g n a l c a n b e m a d e t o s h o w a m u l t i l i n e f e a t u r e , w h i c h suggests t h a t it is also p a r t o f a c l u s t e r o f m a n g a n e s e (34). E x t e n d e d X - r a y a b s o r p t i o n fine s t r u c t u r e ( E X A F S ) studies have b e e n p e r f o r m e d b y various groups o n n u m e r o u s S states o f t h e e n z y m e . T h i s m e t h o d y i e l d s d e t a i l e d i n f o r m a t i o n a b o u t t h e first c o o r d i n a t i o n s p h e r e o f m a n g a n e s e . It is g e n e r a l l y a g r e e d t h a t at least t w o M n - M n v e c t o r s at 2 . 7 Â a r e p r e s e n t as w e l l as a 3 . 3 - Â v e c t o r c o r r e s p o n d i n g to a M n - M ( M n or Ca) i n t e r a c t i o n i n Si. A l s o , the M n atoms are b o u n d to e i t h e r o x y g e n or n i t r o g e n donors w i t h distances


10.


PECORARO

Table I.


Spin

Summary of Conclusions for M n in O E C Manganese Separation

Oxidation State

So Si

Mn^Mn ^ Mn Mn

s

2

Mn

m

s

Mn

I V

3

s

4

Ligands

_

1

m

I V

2

1

Mn

I V

273

2.7 À M n - M n 3.3 À M n - M n or M n - C a 2.7 À M n - M n 3.3 À M n - M n or M n - C a ?

2

3

4

?

Ο, N : carboxylates, imidazole

Ο, N: carboxylates, imidazole, chloride? Ο, N: carboxylates, imidazole, chloride?, water ?

b e t w e e n 1.8 a n d 1.9 À. T h e 2 . 7 - Â d i s t a n c e f u r t h e r i m p l i c a t e s a c l u s t e r of manganese because such close M n - M n separations are o n l y o b s e r v e d i n m o d e l c o m p o u n d s t h a t h a v e s t r u c t u r a l u n i t s s u c h as [ Μ η ( μ - 0 ) ] (36). N o n p r o t e i n - b a s e d l i g a n d s to t h e m a n g a n e s e , i n at least s o m e o f t h e S states, a l m o s t c e r t a i n l y i n c l u d e w a t e r a n d p o s s i b l y c h l o r i d e . T o d e t e r m i n e w h e t h e r n o n e x c h a n g e a b l e w a t e r - d e r i v e d ligands are b o u n d to the M n site p r i o r to t h e o x y g e n e v o l u t i o n s t e p , e x p e r i m e n t s w e r e p e r f o r m e d i n w h i c h t h y l a k o i d m e m b r a n e s are p r e p a r e d i n H O ( i n t h e Si state) and then washed with H O , a f t e r s u b s e q u e n t flashes o f l i g h t . T h e e v o l v e d o x y g e n w a s t h e n c h e c k e d f o r i s o t o p i c c o n t e n t b y mass s p e c t r o m e t r y . I n cases i n w h i c h t h e H O w a s h was d o n e after e i t h e r one or two preflashes, no 0 w a s f o u n d i n t h e e v o l v e d o x y g e n (37). T h i s 2

2

2

s

i e

2

1 8

l

2

i e

2

i m p l i e s that partially o x i d i z e d i n t e r m e d i a t e s are not f o r m e d p r i o r to the S4 S + 0 conversion. T h e reader s h o u l d be aware that this exper i m e n t d o e s n o t r u l e o u t w a t e r l i g a n d s b i n d i n g t o m a n g a n e s e at e a r l i e r stages o f t h e c l u s t e r o x i d a t i o n ( i n r a p i d e x c h a n g e w i t h t h e b u l k s o l v e n t ) b u t d o e s suggest that n o o x i d i z e d f o r m s o f w a t e r ( p e r o x i d e o r s u p e r o x i d e ) a r e o b s e r v e d b e f o r e o r at t h e S s t e p . 0

2

3

A l t h o u g h m a n g a n e s e is a l m o s t c e r t a i n l y t h e site o f r e d o x a d v a n c e ment i n the O E C , water oxidation w i l l not occur unless b o t h C a and C I " i o n s a r e p r e s e n t (38). O t h e r n o n p h y s i o l o g i c a l i o n s (e.g., S r and B r " ) c a n s u b s t i t u t e f o r t h e s e i o n s , b u t l e a d t o far l o w e r a c t i v i t i e s . I n t h e absence of C a or C l ~ the manganese ions may still be o x i d i z e d , l e a d i n g t o S-state a d v a n c e m e n t ; h o w e v e r , 0 is n o t f o r m e d . P r o p o s a l s f o r b o t h calcium and chloride b e i n g directly associated w i t h the manganese clus t e r h a v e b e e n a d v a n c e d (39). U n f o r t u n a t e l y , t h e r e is n o c o m p e l l i n g d a t a t o v e r i f y t h e p r o p o s i t i o n s . L i t t l e is k n o w n o f t h e f u n c t i o n o f c a l c i u m a n d c h l o r i d e . Suggestions for the role of c a l c i u m have i n c l u d e d m a i n t a i n i n g structural integrity, modifying the redox potential of the manganese 2 +

2 +

2 +

2


274


cluster, a n d b i n d i n g a n d t h e n activating w a t e r for catalysis. T h e role of c h l o r i d e is e q u a l l y o b s c u r e a n d m a y i n c l u d e s t a b i l i z i n g t h e m a n g a n e s e c l u s t e r , a c t i n g as a n i n n e r s p h e r e e l e c t r o n - t r a n s f e r m e d i a t o r a n d p r o t e c t i n g t h e c l u s t e r f r o m u n w a n t e d o x i d a t i v e s i d e r e a c t i o n s (see s u b s e quent paragraphs). I n h i b i t i o n o f t h e " S " c y c l e has b e e n s h o w n w i t h s u b s t i t u t e d a m i n e s by causing a two-flash delay of the cycle. T h e reaction of N H O H w i t h d a r k - a d a p t e d P S I I s a m p l e s i n t h e Si state l e a d s t o a t w o - f l a s h d e l a y e d Si state (40), w i t h r e d u c e d M n c e n t e r s (41, 42). H i g h e r c o n c e n t r a t i o n s o f N H O H l e a d t o t h e i r r e v e r s i b l e r e d u c t i o n a n d loss o f t h r e e o f f o u r M n ions. T w o of these ions are released c o o p e r a t i v e l y , p r o b a b l y f r o m the same r e a c t i o n site. A n o t h e r b i n d i n g site, p r e s u m a b l y the f e r r o s e m i q u i n o n e a c c e p t o r s i t e , c a n b e h i t b y v e r y h i g h c o n c e n t r a t i o n s (>6 N H O H / P S I I ) a n d c a u s e i r r e v e r s i b l e s t r u c t u r a l c h a n g e (42). H y d r o q u i n o n e c a n also r e d u c e t h e m a n g a n e s e i n a r e v e r s i b l e m a n n e r ; h o w e v e r , r e c e n t e x t e n d e d X - r a y a b s o r p t i o n fine s t r u c t u r e s p e c t r a s h o w t h a t t h e site o f r e d u c t i o n m a y b e d i f f e r e n t f o r h y d r o x y l a m i n e a n d h y d r o q u i n o n e (41, 43). T h e s e t w o r e d u c t a n t s act s y n e r g i s t i c a l l y at l o w c o n c e n t r a t i o n s t o d e a c t i v a t e t h e e n z y m e c o m p l e t e l y . T h e use o f N H N H , a t w o - e l e c t r o n r e d u c t a n t , has also b e e n s h o w n to r e d u c e t h e s y s t e m b y a t w o - f l a s h d e l a y , a l t h o u g h t h i s m e c h a n i s m is less w e l l - u n d e r s t o o d . U n l i k e h y d r o x y l a m i n e , h y d r o q u i n o n e , o r h y d r a z i n e , a m m o n i a has b e e n s h o w n t o i n hibit the e n z y m e i n a nonredox fashion. A m m o n i a w i l l b i n d reversibly t o M n c l u s t e r i n t h e S state o f P S I I l e a d i n g t o a n e w E P R s i g n a l at g « 4 . 1 (44). W h e n o r i e n t e d s a m p l e s o f t h i s p r o t e i n d e r i v a t i v e a r e e x amined, hyperfine structure can be observed. These hyperfine structures suggest t h a t t h i s l o w - f i e l d E P R s i g n a l is a s s o c i a t e d w i t h a c l u s t e r o f m a n g a n e s e i o n s (34). T h a t a m m o n i a b i n d i n g is c o m p e t i t i v e w i t h C l " a n d i n h i b i t s o x y g e n e v o l u t i o n m i g h t s u g g e s t t h a t c h l o r i d e also b i n d s d i r e c t l y to the manganese cluster; h o w e v e r , a m m o n i a a p p a r e n t l y i n h i b i t s the e n z y m e i n a noncompetitive manner versus chloride, m a k i n g i n t e r p r e tations m o r e difficult. 2


2

2

2

2

2

F r a s c h (45, 46) has s h o w n t h a t t h e O E C c a n c a t a l y z e a n a z i d e - i n s e n s i t i v e catalase r e a c t i o n i n t h e d a r k . T h e a c t i v i t y c a n b e d i r e c t l y ass o c i a t e d w i t h t h e O E C b e c a u s e (1) c o m p e t i t i v e i n h i b i t o r s o f w a t e r o x i d a t i o n a r e also c o m p e t i t i v e i n h i b i t o r s o f t h e catalase a c t i v i t y a n d (2) t h e K j f o r w a t e r o x i d a t i o n a n d catalase a c t i v i t y a r e e s s e n t i a l l y i d e n t i c a l . T h e e n z y m e a p p a r e n t l y c y c l e s i n t h i s case b e t w e e n S a n d S . M a n o a n d c o - w o r k e r s (47) s h o w e d t h a t t h e S i / S _ i states a r e also c o m p e t e n t to c a r r y o u t catalase r e a c t i o n s ; h o w e v e r , t h i s r e a c t i o n is h i g h l y p H - d e p e n d e n t . F o r e x a m p l e , at p H 8 . 8 , t h e S state c a n o x i d i z e H 0 t o 0 , b u t S _ i is i n c a p a b l e o f c o m p l e t i n g t h e r e a c t i o n c y c l e ; h o w e v e r , i f t h e p H is l o w e r e d to p H 6 steady-state m e a s u r e m e n t s o f o x y g e n e v o l u t i o n can b e g a t h e r e d . Just as is t h e case w i t h w a t e r o x i d a t i o n , t h e s e catalase r e a c t i o n s 0

x

2

2

2


2

10.

275


PECORARO

are a c c e l e r a t e d b y C a . Steady-state k i n e t i c analyses o f b o t h S _ i / S i a n d S / S cycles support an o r d e r e d m e c h a n i s m w i t h C a b i n d i n g p r i o r to h y d r o g e n p e r o x i d e . T h e M i c h a e l i s c o n s t a n t f o r H 0 ( 1 7 7 m M ) is c o m p a r a b l e to t h e K f o r t h e M n c a t a l a s e . 2 +

0

2 +

2

2

2

m

N u m e r o u s w o r k e r s h a v e s u g g e s t e d t h a t H 0 m a y b e g e n e r a t e d as an intermediate d u r i n g the water oxidation reactions of photosynthesis. The H 0 e x p e r i m e n t s o f R a d m e r (37) s p e a k against t h i s p r o p o s i t i o n , at least f o r t h e l o w e r S states. H o w e v e r , t h e r e is a n a c c u m u l a t i n g b o d y o f e v i d e n c e that i n d i c a t e s that t h e O E C c a n , u n d e r c e r t a i n c i r c u m s t a n c e s , g e n e r a t e H 0 b y l i g h t - d r i v e n r e a c t i o n s . D e p l e t i o n o f c h l o r i d e at p H 7.2 l e a d s to o p t i m a l h y d r o g e n p e r o x i d e p r o d u c t i o n a n d v i r t u a l l y n o 0 e v o l u t i o n . I n c o n t r a s t , at p H 6 d i o x y g e n is t h e e x c l u s i v e p r o d u c t . T h e r a t e o f p e r o x i d e f o r m a t i o n at p H 7.2 is i n v e r s e l y r e l a t e d t o t h e c h l o r i d e c o n c e n t r a t i o n . O n t h e basis o f t h e s e s t u d i e s , F r a s c h (46,48) has p r o p o s e d t h a t o n e r o l e o f c h l o r i d e m a y b e to p r o t e c t t h e O E C f r o m d i s c h a r g i n g its o x i d i z i n g e q u i v a l e n t s at t o o l o w a n S state to f o r m p e r o x i d e r a t h e r than b e f u r t h e r o x i d i z e d to gather sufficient o x i d i z i n g equivalents to c o n v e r t w a t e r t o d i o x y g e n . A d d i t i o n o f c e r t a i n l i p i d a n a l o g u e s (e.g., l a u r o y l c h o l i n e c h l o r i d e ) t o P S I I m e m b r a n e s has also b e e n s h o w n t o i n h i b i t o x y g e n f o r m a t i o n a n d t o i n d u c e t r a n s i e n t p e r o x i d e f o r m a t i o n (49). It is u n c l e a r f r o m t h i s e x p e r i m e n t h o w t h e l a u r o y l c h o l i n e c h l o r i d e affects t h e m a n g a n e s e c l u s t e r a n d i f it causes a n a l t e r n a t i v e r e a c t i o n p a t h w a y to o c c u r f o r p e r o x i d e f o r m a t i o n . 2

2

1

2

8

2

2


2

M n P e r o x i d a s e . T h e m a n g a n e s e p e r o x i d a s e ( M n P ) is o n e o f t h e two k n o w n enzymes capable of the oxidative degradation of lignin, an amorphous, random, aromatic polymer synthesized from p-hydroxycinnamyl alcohol, 4-hydroxy-3-methoxycinnamyl alcohol, and 3,5,-dimethoxy-4-hydroxycinnamyl alcohol precursors by w o o d y plants. B o t h en zymes contain the p r o t o p o r p h y r i n I X h e m e prosthetic g r o u p , similar to the h e m e peroxidases w i t h an L 5 - h i s t i d i n e a n d b o t h use h y d r o g e n p e r o x i d e as a s u b s t r a t e . H o w e v e r , t h e m a n g a n e s e p e r o x i d a s e has a n a b s o l u t e r e q u i r e m e n t f o r M n ( I I ) t o c o m p l e t e its c a t a l y t i c c y c l e (50). T h e X - r a y s t r u c t u r e o f t h i s p r o t e i n has r e c e n t l y a p p e a r e d (51). T h e p r e s e n t l y a c c e p t e d m e c h a n i s m (52) i n v o l v e s t h e o x i d a t i o n o f an Fe(III) p o r p h y r i n b y h y d r o g e n p e r o x i d e to f o r m an ( F e = 0 ) P * a n a l ogous to the p r e v i o u s l y m e n t i o n e d " c o m p o u n d 1 " o f the h e m e catalase. T h i s h i g h l y o x i d i z e d e n z y m e f o r m s u b s e q u e n t l y reacts w i t h a n e q u i v a l e n t of Mn(II) to give " c o m p o u n d 2 , " ( F e = 0 ) P , a n d M n ( I I I ) , w h i c h can diffuse off o f t h e e n z y m e a n d i n t o t h e m e d i u m . T h e r e is l i t t l e r e s t r i c t i o n f o r t h e t y p e o f M n ( I I ) r e q u i r e d i n t h e first r e d u c t i v e s t e p ; h o w e v e r , t h e subsequent r e d u c t i o n o f c o m p o u n d 2 to r e s t i n g e n z y m e r e q u i r e s an M n ( I I ) d i c a r b o x y l a t e o r α - h y d r o x y a c i d c o m p l e x . S t u d i e s suggest t h a t t h e e n z y m e p r e f e r s t h e 1:1 M n ( I I ) o x a l a t e c o m p l e x as s u b s t r a t e . T h e I V

I V


+

276


o x i d i z e d M n ( I I I ) o x a l a t e is o n c e a g a i n r e l e a s e d i n t o t h e m e d i u m , p r e s u m a b l y f o r m i n g [bis(oxalato)Mn(III)]~, to w r e a k o x i d a t i v e damage o n its p o l y m e r i c s u b s t r a t e t h r o u g h t h e i n i t i a t i o n o f p h e n o l i c r a d i c a l s i n t h e lignin superstructure. A l t h o u g h [bis(oxalato)Mn(III)]~ can o n l y b e f o r m e d t r a n s i e n t l y b e cause o f its o x i d a t i v e i n s t a b i l i t y , m o d e l c h e m i s t r y has s h o w n t h a t M n ( I I I ) a n d M n ( I V ) α - h y d r o x y a c i d s (e.g., l a c t i c a c i d ) c a n b e p r e p a r e d as s t a b l e s o l i d s t h a t c a n t h e n b e u s e d to e v a l u a t e l i g n i n d e g r a d a t i v e r e a c t i o n s (53). T h e [ M n ( I I I ) ( l a c t a t e ) ] " a n d r e l a t e d c o m p o u n d s s u c h as [Mn(III) (lactate) ] w i l l oxidatively degrade vanillin acetone through a r a d i c a l m e c h a n i s m to f o r m p y r u v a l d e h y d e a n d v a n i l l i n . T h e r e a c t i o n r e q u i r e s t w o o x i d i z i n g e q u i v a l e n t s p r o v i d e d b y at least t w o M n ( I I I ) c o m p l e x e s . A l t h o u g h [ b i s ( o x a l a t o ) M n ( I I I ) ] ~ is t h o u g h t t o b e t h e n a t u r a l s u b s t r a t e , [bis(malonato)Mn(III)]~ d o e s n o t a p p e a r to c a t a l y z e t h e v a n i l l i n a c e t o n e d e g r a d a t i o n . P r e s u m a b l y t h i s is d u e t o t h e g r e a t e r s t a b i l i t y o f the [bis(malonato)Mn(III)]~ c o m p l e x . Surprisingly, the m o r e h i g h l y ox i d i z e d m a n g a n e s e c o m p l e x [ M n ( I V ) ( a - h y d r o x y b u t y r i c a c i d ) ] ~ also d o e s not o x i d i z e v a n i l l i n acetone. T h i s m a y i n part be d u e to the e x t r e m e w a t e r s e n s i t i v i t y o f t h i s c o m p l e x (54). 2


2

3

+

3

2

M n Ribonucleotide Reductase. Iron-containing ribonucleotide reductases play an important role i n the transfer of genetic material i n m o s t o r g a n i s m s . H o w e v e r , c e r t a i n C o r y n e f o r m b a c t e r i a s u c h as Br embacterium ammoniagenes a n d Micrococcus luteus r e q u i r e m a n g a n e s e f o r g r o w t h a n d specifically for D N A synthesis. A manganese-containing r i b o n u c l e o t i d e r e d u c t a s e w a s i s o l a t e d a n d p u r i f i e d f r o m B. ammoniagenes (55, 56). T h e a c t i v e e n z y m e a p p e a r s t o c o n s i s t o f a d i m e r (B2) o f M W = 1 0 0 , 0 0 0 d a l t o n s a c t i n g as t h e c a t a l y t i c s u b u n i t a l o n g w i t h a n u c l e o t i d e b i n d i n g m o n o m e r ( B l ) o f M W = 8 0 , 0 0 0 d a l t o n s . T h e e n z y m e is b e l i e v e d t o c o n t a i n a d i n u c l e a r a c t i v e s i t e . T h e p r e s e n c e o f M n ( I I I ) is s u g g e s t e d b y the similarity of the o p t i c a l a b s o r p t i o n to the same d i m e r i c M n ( I I I ) m o d e l c o m p l e x e s that have b e e n u s e d i n r e l a t i o n to the manganese cat alases (see M n C a t a l a s e s ) . F u r t h e r m o r e , t h e a c t i v e e n z y m e is E P R s i l e n t , w h i c h is c o n s i s t e n t w i t h e i t h e r M n ( I I I ) o r a s t r o n g l y c o u p l e d M n ( I V ) d i m e r . O n t h e basis o f t h e s e l i m i t e d o b s e r v a t i o n s , it has b e e n s u g g e s t e d that the manganese a n d i r o n r i b o n u c l e o t i d e reductases m a y f u n c t i o n i n analogous ways. T h e site i n t h e a c t i v e F e r i b o n u c l e o t i d e r e d u c t a s e c o n t a i n s t w o F e ( I I I ) i o n s 3.3 À a p a r t , b r i d g e d b y o n e c a r b o x y l a t e f r o m a g l u t a m a t e r e s i d u e a n d a w a t e r - d e r i v e d o x o b r i d g e (57). T h e f u n c t i o n o f t h i s i r o n c e n t e r appears to b e the f o r m a t i o n a n d s t a b i l i z a t i o n o f a free r a d i c a l o n a t y r o s i n e a b o u t 5 À a w a y . T h i s r a d i c a l is f o r m e d b y r e a c t i o n o f t h e reduced, diferrous center w i t h 0 , probably through peroxide and ferryl i n t e r m e d i a t e s . T h i s u n u s u a l l y s t a b l e t y r o s y l r a d i c a l is t h o u g h t t o p a r t i e 2


10.

PECORARO


277

ipate i n the r e d u c t i o n of ribonucleotides b y electron transfer o f the free radical, possibly t h r o u g h t h e F e center, to t h e substrate interaction sur f a c e i n w h i c h t h e r i b o s e r i n g is o x i d i z e d b y o n e e l e c t r o n t o a c t i v a t e i t


f o r r e a c t i o n w i t h r e d o x - a c t i v e t h i o l s i n t h e p r o t e i n (58). M n Superoxide Dismutase. T h e manganese superoxide dismutase ( S O D ) contains a single M n i o n that shuttles b e t w e e n t h e +2 a n d + 3 o x i d a t i o n l e v e l d u r i n g t h e c a t a l y t i c c y c l e . T h e M n S O D is t h e b e s t u n d e r s t o o d o f a l l t h e m a n g a n e s e r e d o x e n z y m e s . T h i s is p a r t i a l l y due to the high-resolution X - r a y structures o f the Mn(II) a n d Mn(III) e n z y m e s f r o m Thermus thermophilus (59, 60), t h e d e t a i l e d k i n e t i c a n a l ysis o f t h e e n z y m e that h a s b e e n u n d e r t a k e n (61, 62), a n d ( o n c e again) the similarity of this manganoenzyme to an i r o n e n z y m e o f like function (63). T h e X - r a y c r y s t a l s t r u c t u r e s o f t h e M n a n d F e S O D s i n d i c a t e t h a t these t w o proteins are v e r y similar, b o t h i n terms o f the active-site ge o m e t r y a n d a h i g h d e g r e e o f s e q u e n c e h o m o l o g y (63). B o t h t h e m a j o r global fold a n d the metal-active center are essentially unaltered w h e n t h e e n z y m e is r e d u c e d t o t h e M n ( I I ) f o r m . T h e m e t a l i o n i n b o t h t h e M n a n d F e S O D s h a v e a t r i g o n a l b i p y r a m i d a l l i g a n d a r r a y , as s h o w n i n F i g u r e 3, w i t h t w o h i s t i d i n e s a n d a c a r b o x y l a t e i n t h e e q u a t o r i a l p o s i t i o n s and an additional histidine and a solvent-derived hydroxide or water i n the axial positions. A crystal structure o f t h e a z i d e - b o u n d derivative o f F e ( S O D ) shows that small m o l e c u l e s m a y b i n d to t h e active-site m e t a l m a k i n g i t s i x - c o o r d i n a t e (64). T h i s suggests d i r e c t b i n d i n g o f s u p e r o x i d e to t h e m e t a l site d u r i n g t h e c a t a l y t i c d i s p r o p o r t i o n c y c l e .

(GLU) Ο H N ^ r '

(HIS)

Κ

NH '(HIS)

(Hisy Figure 3. The active site of Mn superoxide dismutase on the basis of the X-ray structure ofLudwig et al. (59). The "W" represents a water-derived ligand, either OH~ or H 0. 2


278


K i n e t i c studies of the F e ( S O D ) indicate a fairly simple o x i d a t i o n r e d u c t i o n c y c l e i n w h i c h 0 ~ is b o u n d t o t h e F e ( I I I ) f o r m o f t h e p r o t e i n a n d o x i d i z e d to 0 , f o l l o w e d b y b i n d i n g o f a s e c o n d 0 " t o t h e r e s u l t i n g Fe(II) f o r m a n d r e d u c t i o n to H 0 . T h e k i n e t i c s o f M n ( S O D ) are m o r e c o m p l i c a t e d a n d t h e t u r n o v e r n u m b e r ( 1 3 0 0 s " at 2 5 ° C ) is m u c h l o w e r t h a n f o r F e ( S O D ) ( 2 6 , 0 0 0 s " ) ; h o w e v e r , a s i m i l a r c a t a l y t i c c y c l e is b e l i e v e d to o c c u r . T h e manganese e n z y m e k i n e t i c s are c o m p l i c a t e d b y a side r e a c t i o n to f o r m a " d e a d e n d c o m p l e x , " possibly a Mn(III) p e r o x i d e c o m p l e x (61, 62). 2

2

2

2

2

1


1

Biomimetic Reactions Using Synthetic Model Compounds T h e research groups interested i n the reaction chemistry of manganese have focused on understanding either the assembly reactions of m a n ganese clusters or the reactivity o f manganese c o m p o u n d s w i t h H 0 , H 0 , H 0 , 0 , or iodosylbenzene. In the majority of the cluster assem b l y r e a c t i o n s o n e is e x a m i n i n g a c i d - b a s e s u b s t i t u t i o n r e a c t i o n s t h a t i n t e r c o n v e r t stable or meta-stable aggregations of manganese oxo cores. W e w i l l not d w e l l o n these reactions i n this section b u t rather direct t h e i n t e r e s t e d r e a d e r to t h e l i t e r a t u r e i n t h i s a r e a (65-67). M o r e p e r t i n e n t t o t h e p o i n t o f e n z y m e m i m e t i c c h e m i s t r y is t h e r e a c t i v i t y o f small molecules w i t h Mn(II) through M n ( V ) . T h e following section de s c r i b e s t h e w o r k t h a t w e h a v e d o n e i n o u r l a b o r a t o r y as a n e x a m p l e o f h o w t h e s t u d y o f i n o r g a n i c m o d e l c o m p l e x e s c a n l e a d to a n u n d e r s t a n d i n g of possible mechanisms for the r e a c t i v i t y f o u n d i n b i o l o g i c a l systems. F o r m o r e d e t a i l s o n w o r k d o n e b y o t h e r g r o u p s i n t h i s field, t h e r e a d e r is d i r e c t e d t o a r e c e n t r e v i e w o n t h i s c h e m i s t r y (68). 2

2

2

2

2

Models for the Reactivity of OR)] Cores in Photosynthesis.

[ Μ η ( ΐ ν ) ( μ - 0 ) ] and [ Μ η ( Ι Ι Ι ) ( μ A s discussed i n the previous para 2 g r a p h s , t h e o x y g e n - e v o l v i n g c o m p l e x has a n a s s e m b l y o f f o u r m a n g a n e s e ions that f o r m the heart of the w a t e r oxidation reaction. T h e manganese ions appear to b e a r r a n g e d i n groups o f [ Μ η ( μ - 0 ) ] units ( M n - M n sep a r a t i o n « 2.7 Â ) w i t h t h e o x i d a t i o n state o f t h e m a n g a n e s e v a r y i n g between Mn(III) and M n ( I V ) depending on the enzyme oxidation level ( S states). I n S , t h e e n z y m e is b e l i e v e d t o c o n t a i n 1 M n ( I I I ) a n d 3 M n ( I V ) i o n s (23). T h i s state c a n t u r n o v e r c a t a l y t i c a l l y w i t h So, [ M n ( I I I ) M n ( I V ) i], i n a catalase r e a c t i o n (45, 48). O n t h e basis o f t h e s e observations, w e b e l i e v e d that it was i m p o r t a n t to u n d e r s t a n d h o w [Μη(ΐν)(μ -0)] c o u l d be prepared and h o w they behave with respect t o catalase r e a c t i v i t y . 2

2

n

2

2

2

2

3

2

2

O u r first task w a s to u n d e r s t a n d p r e c i s e l y h o w M n ( I I I ) d i m e r s c o u l d b e c o n v e r t e d to [ Μ η ( ΐ ν ) ( μ - 0 ) ] c o r e s . W e p r e p a r e d [ M n ( I I I ) ( 3 , 5 - d i C l s a l p n ) ^ - O C H ) ] ( s a l p n is l , 3 - b i s ( s a l i c y l i d e n e i m a n a t o ) p r o p a n e ) , t h e 2

2

3

2

2


10.

279


PECORARO

first e x a m p l e o f a d i a l k o x i d e - b r i d g e d M n ( I I I ) d i m e r h a v i n g J a h n - T e l l e r d i s t o r t i o n s a l o n g t w o o f t h e M n - O R b o n d s (36). T h e t e t r a d e n t a t e s a l p n l i g a n d a d o p t e d t h e cis-β c o n f i g u r a t i o n i n t h i s i s o m e r . T h i s m o l e c u l e is air-sensitive, c o n v e r t i n g after several h o u r s to [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] . T h i s [ Μ η ( ΐ ν ) ( μ - 0 ) ] d i m e r has a M n - M n s e p a r a t i o n o f 2 . 7 2 À , w h e r e a s i n t h e [ Μ η ( Η Ι ) ( μ - Ο Κ ) ] d i m e r t h e s e p a r a t i o n is 3 . 1 9 À (36, 69). T h e s e m o l e c u l e s , w h i c h f o r m t h e basis o f t h i s s t u d y , a r e s h o w n i n F i g u r e 4. 2

2

2

2

Formation of [Mn(IV)(salpn)^ -0)] Using Hydrogen Peroxide. T h e c o m p l e x [ M n ( I I I ) (salpn) ( C H 0 H ) ] C 1 0 is u n r e a c t i v e w i t h H 0 w h e n dissolved i n methanol, D M F , or acetonitrile; h o w e v e r , add i t i o n o f a b a s e s u c h as N a O H o r N a O M e l e a d s t o r a p i d f o r m a tion of [Mn(IV)(salpn)( -0)] (70). W e s h o w e d t h a t [Μη(III)(salpn) ( C H 0 H ) ] C 1 0 c o n v e r t s t o [ M n ( I I I ) (salpn) ( μ - Ο Ο Η ) ] i f N a O M e is a d d e d to a d e g a s s e d a c e t o n i t r i l e s o l u t i o n . R e a c t i o n o f t h i s M n ( I I I ) d i m e r w i t h p e r o x i d e i n a c e t o n i t r i l e is r a p i d a n d q u a n t i t a t i v e i n t h e p r o duction of [Mn(IV)(salpn)^ -0)] . D i o x y g e n w i l l also r e a c t w i t h [ M n ( I I I ) ( s a l p n ) ( M - O C H ) ] to give [ M n ( I V ) ( s a l p n ) 0 ] ; h o w e v e r , the r e a c t i o n is ~ 1 0 0 0 t i m e s s l o w e r a n d g i v e s « 7 0 % p r o d u c t . M a s l e n a n d W a t e r s (71) c h a r a c t e r i z e d t h e p r o d u c t o f d i o x y g e n r e a c t i o n w i t h Μη(II)(salpn) b y X - r a y c r y s t a l l o g r a p h y . T h e y i n t e r p r e t e d i t as [Mn(III)(salpn)^ -OH)] because Mn(IV) complexes were considered u n l i k e l y at t h e t i m e . B o u c h e r a n d C o e (72) s u g g e s t e d a [ Μ η ( ΐ ν ) ( μ - 0 ) ] f o r m u l a t i o n f o r t h a t s t r u c t u r e a n d t h a t t h e p r o d u c t w a s t h e s a m e as f o r t h e o x i d a t i o n o f [ M n ( I I I ) ( s a l p n ) ] . T h e y w e r e s h o w n b y o u r s t u d i e s (36, 70) a n d A r m s t r o n g ' s (69) t o b e c o r r e c t . 2

2

3

2


2

2

2

4

2

M 2

3

2

2

4

2

2

2

3

2

3

2

2

2

2

2

2

2

+

A n important question i n the [Mn(IV)(salpn)(μ -0)] formation re a c t i o n is " w h i c h m o l e c u l e ( s ) is(are) t h e s o u r c e o f t h e b r i d g i n g o x i d e s i n the M n ( I V ) d i m e r ? W e addressed this question b y f o l l o w i n g the i n corporation of O from labeled 0 , H 0 , and water into [Mn(IV)( s a l p n ) ^ - 0 ) ] (69). T h e m o l e c u l a r i o n i n t h e n e g a t i v e i o n F A B mass spectrum of [Mn(IV)(salpn)(μ -0)] a l l o w e d for d e t e c t i o n a n d q u a n t i tation of labeled product [Mn(IV)(salpn)( - 0)] , 702; [Mn(IV) (salpn) ( - 0, μ - 0 ) ] , 704; [Mn(IV)(salpn)( - 0)] , 706 amu. D r o p wise addition of —0.1 M H 0 in H O to an a c e t o n i t r i l e s o l u t i o n o f [ M n ( I I I ) ( s a l p n ) ^ - O C H ) ] resulted exclusively i n the doubly labeled [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] . Similar results w e r e obtained u n d e r aerobic ( 0 ) a n d a n a e r o b i c c o n d i t i o n s . N o r i s e is o b s e r v e d a b o v e t h e s t a t i s t i c a l d i s t r i b u t i o n o f t h e mass p e a k at 7 0 4 i n s p i t e o f a n 5 5 0 - f o l d m o l a r e x c e s s of H 0 . T h i s c l e a r l y d e m o n s t r a t e s that t h e p r i m a r y r e a c t i o n m e c h a n i s m excludes oxygen contained in water from entering the bridges i n [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] . T o test w h e t h e r b o t h o x i d e a t o m s a r e g e n erated from the same m o l e c u l e of h y d r o g e n p e r o x i d e , w e e x a m i n e d m i x t u r e s of 0.1 M aqueous ( H O ) H 0 , and H 0 with 2

2

, ,

l s

2

2

2

2

2

2

2

1 6

M 2

2

M 2

1 6

2

1 8

M 2

2

2

2

1 8

3

1 8

1

8

2

2

2

1 8

2

2

i e

2

2

2

2

1 6

2

2

2

i e

2

1

6

2

2

1

8


2


Figure 4. (A) ORTEP diagram of[Mn(IV) (salpn) (μ -0)] . gram of [Mn(III) (salpn) (μ -Ο0Η )] . 2

2

3

2

(Β) ORTEP dia

2


10.

281


PECORARO

[ M n ( I I I ) ( s a l p n ) ( C H 0 ) ] . T h e p r i m a r y mass p e a k s a r e o b s e r v e d at 7 0 2 and 706 amu, respectively, w i t h no observed increase i n the m i x e d d i m e r mass p e a k at 7 0 4 . F r o m t h e s e r e s u l t s w e c o n c l u d e t h a t h y d r o g e n p e r o x i d e is t h e s o u r c e o f t h e o x i d e o x y g e n s i n [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] a n d that b o t h o x i d e - b r i d g i n g atoms originate f r o m the same p e r o x i d e m o l ecule. This isotopic distribution demonstrates that m o n o m e r i c inter 3

2

2

2

m e d i a t e s s u c h as L M n ( I V ) 0 o r L M n ( V ) 0 a r e n o t p a r t o f t h e r e a c t i o n m e c h a n i s m i n t h i s case. A l t h o u g h h i g h - v a l e n t , m o n o m e r i c complexes that m i g h t f o r m after p e r o x i d e b o n d b r e a k i n g h a v e b e e n e l i m i n a t e d as i n t e r m e d i a t e s i n t h e formation of [Mn(IV)(salpn)(^ -0)] , the existence of a m o n o m e r i c i n t e r m e d i a t e p r i o r t o h y d r o g e n p e r o x i d e c l e a v a g e m u s t also b e a d d r e s s e d . T h i s point was tested b y r e a c t i n g a 5 0 : 5 0 m i x t u r e o f [Mn(III)(salpn)(μ O C H ) ] and [Mn(III)(3,5-diCl-salpn)(M -OCH )] w i t h H 0 i n aceto n i t r i l e . I n this case, a m i x t u r e o f [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] , [ M n ( I V ) ( 3 , 5 diCl-salpn)(salpn)(M -0)], a n d [Mn(IV)(0)(3,5-diCl-salpn)(M -0)] was obtained. W h e n mixed, neither [Mn(IV)(salpn)^ -0)] and [Mn(IV)(3,5d i C l - s a l p n ) ( M - 0 ) ] n o r [Mn(III) (salpn) ( - O C H ) ] a n d [ M n ( I I I ) ( 3 , 5 - d i C l s a l p n ) ^ - O C H ) ] w i l l s c r a m b l e o n t h e t i m e scale o f t h e e x p e r i m e n t t o give [Mn(III) (3,5-diCl-salpn)(salpn)(M -OCH ) ] or [ M n ( I V ) ( 3 , 5 - d i C l salpn) (salpn) ( μ - 0 ) ] , r e s p e c t i v e l y . T h u s , i n s e r t i o n o f h y d r o g e n p e r o x i d e requires the dissociation of the Mn(III) d i m e r p r i o r to cleavage of the peroxide bond.


2

2

2

3

2

2

3

2

2

2

2

2

2

2

3

2

2

2

2

2

M 2

3

2

2

2

2

2

2

2

3

2

2

2

A n o t h e r i m p o r t a n t consideration for the evaluation of reactivity i n v o l v i n g f o r m a l o x i d a t i o n state c h a n g e s is t h e e l e c t r o c h e m i c a l p o t e n t i a l of the reactants. T h e m o n o m e r i c [ M n ( I I I ) ( s a l p n ) ( C H O H ) ] C 1 0 shows an irreversible one-electron r e d u c t i o n (Mn(III) Mn(II), confirmed by r o t a t i n g p l a t i n u m e l e c t r o d e v o l t a m m e t r y , i n a c e t o n i t r i l e at — 1 0 6 m V versus S C E . W e have o b s e r v e d no oxidative e l e c t r o c h e m i s t r y for this c o m p o u n d out to potentials of +1 V . I n contrast, [Μη(III)(salpn)(μ O C H ) ] show a quasireversible, one-electron oxidation around + 5 5 0 m V a n d an i r r e v e r s i b l e o n e - e l e c t r o n r e d u c t i o n a r o u n d —650 m V . T h e d r a m a t i c s t a b i l i z a t i o n o f t h e M n ( I V ) o x i d a t i o n l e v e l is l i k e l y a r e s u l t o f the additional basic oxyanion per manganese, w h i c h provides another negative charge to the M n i o n . 3

4

2

3

2

A Proposed M e c h a n i s m for the F o r m a t i o n of [Mn(IV)(salpn)^ -0)] . I f h y d r o g e n p e r o x i d e is a d d e d t o [ M n ( I I I ) ( s a l p n ) ( μ O C H ) ] i n degassed a c e t o n i t r i l e , an instantaneous r e a c t i o n occurs to give [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] i n > 9 7 % y i e l d . A mechanistic proposal for t h e p r o d u c t i o n o f [ M n ( I V ) (salpn) ( μ - 0 ) ] b y r e a c t i o n o f H 0 with [ M n ( I I I ) ( s a l p n ) ^ - O C H ) ] is s h o w n i n F i g u r e 5. T h i s s c h e m e is c o n s i s t e n t w i t h b o t h t h e r e a c t i v i t y a n d i s o t o p i c - l a b e l i n g s t u d i e s p r e s e n t e d . It f e a t u r e s H 0 r e a c t i n g v i a t w o s u c c e s s i v e d e p r o t o n a t i o n steps t h a t u s e 2

3

2

2

2

2

2

2

2

2

3

2

2

2

2


2

282



2, X-ray

Figure 5. The proposed mechanism for the formation of [Mn(IV) (salpn)(μ -0)\ from [Mn(III)(salpn) (p -OCH )] and hydrogen peroxide. 2

2

2

3

2


10.

283


PECORARO

basic oxyanions associated w i t h the Mn(III) precursors. T h e

presence

o f s o m e f o r m o f b a s e is a n a b s o l u t e r e q u i r e m e n t as i l l u s t r a t e d b y t h e differential

reactivity

[ M n ( I I I ) (salpn) ( μ - Ο Ο Η ) ] 2

3

of 2

[Mn(III)(salpn)(CH OH)]Cl0 3

versus

4

(69). A l s o , t h e Ή N M R a n d e l e c t r o c h e m i c a l

studies u s i n g t r i e t h y l a m i n e i n d i c a t e that the abilities of oxyanions to s t a b i l i z e t h e M n ( I V ) o x i d a t i o n l e v e l o r to p r o m o t e f o r m a t i o n o f " p r e t e m p l a t e d " complexes

cis-β

are not the necessary characteristics that

i m p a r t r e a c t i v i t y . W e c o n c l u d e t h a t it is t h e p r o t o n - a c c e p t i n g

feature

o f t h e s e a n i o n s t h a t a l l o w s o x i d a t i o n at t h e m e t a l c e n t e r a n d t h a t d e -


protonation must, therefore, occur before cleavage bond. Although H 0 2

may transiently coordinate

2

of the to

peroxide

[Mn(III)(salpn)

( C H 0 H ) ] C 1 0 , m e t a l - c e n t e r e d oxidation p r o b a b l y does not occur p r i o r 3

4

to d e p r o t o n a t i o n . It is n o t a b l e t h a t a s s o c i a t i o n o f t h e

hydroperoxide

a n i o n to M n ( I I I ) c r e a t e s a l i g a n d e n v i r o n m e n t s i m i l a r to t h a t s e e n i n [ M n ( I I I ) ( s a l p n ) ^ - O C H ) ] . T h u s , d e p r o t o n a t i o n leads to stabilization 2

3

2

o f t h e M n ( I V ) o x i d a t i o n l e v e l at t h e m e t a l c e n t e r a n d r e p l a c e m e n t LM

3

for H

+

on H 0

+

2

2

(giving [ L M

3 +

of

( 0 H ) ] ) may make the coordinated 2

p e r o x i d e m o l e c u l e m o r e easily r e d u c e d . B o t h o f these factors s h o u l d l e a d to an increase i n the d r i v i n g force for i n t e r n a l o x i d a t i o n o f the i n t e r m e d i a t e . O u r o b s e r v a t i o n s a r g u e f o r d i s s o c i a t i o n t h a t is i n s t i g a t e d by H 0 2

2

rather than a predissociation step. W e prefer the f o r m u l a t i o n

o f 5 o r 6 c o n v e r t i n g t o 7 as t h e i n t e r m e d i a t e s i n t h i s p r o c e s s ;

however,

o n t h e basis o f t h e s e d a t a , w e c a n n o t e x c l u d e a S t o m b e r g - t y p e p e r o x o m o n o m e r (e.g., [ M n ( I I I ) ( s a l p n ) ( 0 ) ] ~ 2

as s e e n f o r V ( V ) c o m p l e x e s ) t h a t

c o u l d t h e n r e a c t w i t h a m o n o m e r i c M n ( s a l p n ) t o f o r m [Mn(IV)(salpn)(μ +

0)] .

A

2

pathway

as s h o w n

in Figure 5

2

is c o n s i s t e n t

with

these

observations. T h e f o r m a t i o n o f [ M n ( I V ) (salpn) ( μ - 0 ) ] 2

OCH )] 3

2

2

from

[Mn(III)(salpn)(μ 2

u s i n g f - b u t y l h y d r o p e r o x i d e as t h e o x i d a n t a p p e a r s t o f o l l o w a

p a t h w a y v e r y d i f f e r e n t f r o m that o f r e a c t i o n w i t h H 0 2

2

(69). T h e r e a c t i o n

is n o t n e a r l y as fast, r e q u i r i n g a n u m b e r o f h o u r s (at r o o m t e m p e r a t u r e ) and the presence of water. f - B u t y l h y d r o p e r o x i d e lacks a second dis sociable p r o t o n . T o f o r m [Mn(IV)(salpn)(μ -0)] , it c o u l d lose f - b u t y l 2

2

cation, l e a d i n g to conservation of b o t h u n l a b e l e d p e r o x i d e oxygens i n [Mn(IV)(salpn)^ -0)] . 2

2

H o w e v e r , the operative m e c h a n i s m allows ex

t e n s i v e m i x i n g o f l a b e l e d w a t e r . T h e i s o t o p i c p a t t e r n is c o n s i s t e n t w i t h the initial f o r m a t i o n of M n ( V ) ( s a l p n ) 0 , w h i c h t h e n reacts w i t h an e q u i v alent o f M n ( I I I ) s a l p n ( O H ) to give [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] , 2

2

although n u

m e r o u s p a t h w a y s c o u l d b e e n v i s i o n e d t h a t m i g h t fit t h e s e d a t a . F i n a l l y , the reaction of [Mn(III)(salpn)(μ -ΟΟΗ )] 2

3

2

with dioxygen

apparently

f o l l o w s a d i f f e r e n t (or at least a d d i t i o n a l ) p a t h w a y b e c a u s e d i r e c t o x i dation by

1 6

0

2

i n the presence

of a small amount of

1 8

O H ~ leads to

s c r a m b l i n g o f t h e l a b e l i n t h e o x o b r i d g e s . T h i s is i n c o n t r a s t t o t h e


284


h y d r o g e n p e r o x i d e oxidations for w h i c h l a b e l i n g of the b r i d g i n g oxides was not d e p e n d e n t o n the presence of e i t h e r H 0 or OH". 1

2

1 8

Catalase Reactions Using [Mn(IV)(salpn)(M 2 -0)] 2 . T h e r e a c tions d e s c r i b e d p r e v i o u s l y investigated the m e c h a n i s m of formation of [ Μ η ( ΐ ν ) ( μ - 0 ) ] units that h a d structures a p p r o p r i a t e to m o d e l a single d i m e r i n the " d i m e r of dinners" structural proposal for the O E C . T h e o b s e r v a t i o n t h a t t h e O E C c o u l d u n d e r g o a catalase r e a c t i o n b y u s i n g s u c h M n ( I V ) d i m e r s i n s p i r e d us t o e x a m i n e t h e r e a c t i v i t y o f [Mn(IV)(salpn)(μ -0)] w i t h h y d r o g e n p e r o x i d e . T h e o b s e r v e d catalase reactions were c o m p l e t e d b y the addition of a small aliquot of « 5 m M H 0 i n a c e t o n i t r i l e to a d i c h l o r o m e t h a n e s o l u t i o n c o n t a i n i n g [ M n ( I V ) (salpn) ( μ - 0 ) ] . T u r n o v e r o f h y d r o g e n p e r o x i d e to y i e l d d i o x y g e n w a s m o n i t o r e d b y m a n o m e t r y a n d s h o w n t o b e q u a n t i t a t i v e i n less t h a n 1 m i n u s i n g as m u c h as 1 0 0 0 - f o l d m o l a r excess H 0 . F u r t h e r m o r e , greater than 9 8 % of the starting catalyst was r e c o v e r e d . T h e s e obser vations clearly d e m o n s t r a t e d that [Mn(IV)(salpn)(μ -0)] was an effective " c a t a l a s e " (73). 2

2

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A t this p o i n t , w e a p p l i e d o u r i s o t o p e - l a b e l i n g m e t h o d o l o g y to an e x a m i n a t i o n o f t h i s catalase r e a c t i o n . T h e r e a c t i o n o f [Μη (IV) (salpn) ( μ 0 ) ] with H 0 y i e l d e d exclusively 0 , whereas the reverse c o m bination [Mn(IV)(salpn)( - 0)] and H 0 gave the expected 0 . T h i s d e m o n s t r a t e s t h a t d i o x y g e n is d e r i v e d e x c l u s i v e l y f r o m h y d r o g e n p e r o x i d e and not f r o m the μ -οχο linkages of the d i m e r . T h e reaction of [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] w i t h a 1:1 m i x t u r e o f H 0 and H 0 gave a m i x t u r e of 0 a n d 0 w i t h o n l y the p r e d i c t e d statistical d i s t r i b u t i o n of 0 based on residual H 0 . T h u s , b o t h o x y g e n atoms of d i o x y g e n must c o m e f r o m the same h y d r o g e n p e r o x i d e m o l e c u l e . T h i s iso t o p e - l a b e l i n g p a t t e r n is i d e n t i c a l to t h a t r e p o r t e d f o r t h e L. plantarum M n catalase a n d t h e o x y g e n - e v o l v i n g c o m p l e x . T h e i s o t o p i c c o m p o s i t i o n of [Mn(IV)(salpn)(μ -0)] r e c o v e r e d f r o m the catalase e x p e r i m e n t s showed substantial enrichment of label into the μ - 0 " . Single turnover r e a c t i o n s c o m p l e t e d u n d e r h i g h d i l u t i o n c o n d i t i o n s u s i n g a 2:1 r a t i o of H 0 and [Mn(IV)(salpn)^ - 0)] gave p r e d o m i n a n t l y (>95%) [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] , d e m o n s t r a t i n g that the μ - 0 ligands are stoichiometrically exchanged d u r i n g the course of the reaction. T h e re a c t i o n o f t h e 1:1 m i x t u r e o f H 0 :H 0 i n a t w o f o l d excess o v e r [Mn(IV)(salpn)( - 0)] gives [ M n ( I V ) ( s a l p n ) ( - 0 ) ] and [Mn(IV)( s a l p n ) ( ^ - 0 ) ] i n e q u a l r a t i o s , b u t n o i n c r e a s e i n [Μη (IV) (salpn) ( μ 0 ) ] . T h e s e e x p e r i m e n t s s t r o n g l y suggest t h a t t h e c a t a l a s e r e a c t i o n results i n b o t h b r i d g i n g oxo g r o u p s o r i g i n a t i n g f r o m the same p e r o x i d e m o l e c u l e . T h e r e a c t i o n o f a 1:1 m i x t u r e o f [ M n ( I V ) ( s a l p n ) ( μ - 0 ) ] and [ M n ( I V ) ( 3 , 5 - d i C l - s a l p n ) ^ - 0 ) ] w i t h h y d r o g e n p e r o x i d e gave a m i x ture of [Mn(IV)(salpn)( -0)] , [Mn(IV) (3,5-diCl-salpn)(salpn)(M -0)] 2

1 6

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PECORARO

a n d [ M n ( I V ) ( 3 , 5 - d i C l - s a l p n ) ^ - 0 ) ] . A 1:1 m i x t u r e o f t h e p a r e n t [ M n ( I V ) ( s a l p n ) ( M - 0 ) ] a n d [ M n ( I V ) ( 3 , 5 - d i C l - s a l p n ) ( - 0 ) ] is s t a b l e t o ligand exchange u n d e r these conditions i n the absence o f h y d r o g e n p e r o x i d e . T h e r e f o r e , s c r a m b l i n g o f t h e M n L u n i t s r e q u i r e s that a m o n o m e r i c intermediate must b e f o r m e d d u r i n g the reaction. T h e s e o b s e r v a t i o n s s u g g e s t e d t h a t t h e first r e d o x s t e p o f t h e p r o c e s s was initial oxidation o f h y d r o g e n p e r o x i d e to give a Mn(III) species. T h e electrochemistry of [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] shows reversible, one-elec t r o n r e d u c t i v e e l e c t r o c h e m i s t r y at - 4 8 0 m V v e r s u s S C E t o f o r m [ M n ( I I I / I V ) ( s a l p n ) ^ - 0 ) ] . A second reduction wave was not observed. Protonation of [Mn(IV)(salpn)^ -0)] at t h e o x o b r i d g e t o f o r m [ Μ η ( ΐ ν ) ^ 1 ρ η ) ( μ - 0 ) ( μ - Ο Η ) ] c a n also b e a c h i e v e d . T h e e l e c t r o c h e m i s t r y o f t h i s s p e c i e s is less r e v e r s i b l e t h a n f o r [ M n ( I V ) ( s a l p n ) ( μ 0 ) ] a n d occurs nearly 8 0 0 m V to m o r e positive potential, consistent w i t h a decrease i n donation of the b r i d g i n g ligand. Analysis of E X A F S spectra o f [Μη(ΐν) ^1ρη) (μ -0)(μ -ΟΗ)] r e v e a l that t h e M n - M n sep a r a t i o n i n c r e a s e s b y 0.1 À t o 2 . 8 1 À , a g a i n c o n s i s t e n t w i t h a p o o r e r d o n a t i n g b r i d g i n g l i g a n d (72). I n a d d i t i o n , [ Μ η ( ΐ ν ) ^ 1 ρ η ) ( μ - 0 ) ( μ O H ) ] is n o t c o m p e t e n t i n t h e catalase r e a c t i o n w i t h H 0 b u t w i l l r e a c t w i t h N a H 0 (72). T h e s e o b s e r v a t i o n s suggest t h a t p r o t o n s m a y b e v e r y i m p o r t a n t i n t h e s e t y p e s o f c a t a l y t i c r e a c t i o n s . I n fact, B o u c h e r a n d C o e (72) c l a i m t o h a v e d e t e c t e d h y d r o g e n p e r o x i d e p r o d u c e d i n s o l u t i o n s of the Schiff base c o m p l e x , [Μη(ΐν)(Βυ^1ρη)(μ -0)] , u p o n a d d i t i o n o f H C 1 0 . T h e y suggest that t h i s h y d r o g e n p e r o x i d e p r o d u c t i o n o c c u r s b y p r o t o n a t i o n o f b o t h oxo b r i d g e s , f o l l o w e d b y d e c o m p o s i t i o n to H 0 a n d M n ( I I I ) c o m p l e x e s . A p y r o g a l l o l s o l u t i o n w a s u s e d t o test f o r o x y g e n evolution u p o n addition of acid; however, none was detected. 2

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It is w o r t h c o n s i d e r i n g w h y [ M n ( I V ) ( s a l p n ) ( μ - 0 ) ] c a n b e p r o t o n a t e d a n d is a n e f f i c i e n t c a t a l a s e , w h e r e a s p r e v i o u s [ Μ η ( ΐ ν ) ( μ - 0 ) ] c o m p l e x e s d o n o t s h o w s i m i l a r c h e m i s t r y . I n m o s t cases, t h e [ Μ η ( ΐ ν ) ( μ - 0 ) ] c o r e is f o u n d i n s y s t e m s that c o n t a i n n e u t r a l n i t r o g e n d o n o r l i g a n d s . R e c e n t e l e c t r o c h e m i c a l studies of such c o m p o u n d s have s h o w n p K a values o f « 11 f o r [Μη(ΙΙΙ)(μ -0)] a n d « 2 f o r Μ η ( Ι Ι Ι ) Μ η ( ΐ ν ) ( μ - 0 ) ( 7 5 , 76). T h e anionic charge a n d stronger donor ability o f phenolates c o m p a r e d to p y r i d i n e a n d r e l a t e d ligands leads to m o r e e l e c t r o n density o n t h e m e t a l c e n t e r s . T h e r e f o r e , t h e o x o b r i d g e s a r e n o t r e q u i r e d t o d o n a t e as m u c h electron density to the metal center a n d can b e i n v o l v e d i n p r o t o n ac c e p t o r c h e m i s t r y . N o r t o n (77) s h o w e d t h a t p r o t o n a t i o n rates o f m o l e c u l e s s u c h as [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] a r e r e l a t i v e l y s l o w . T h i s a p p e a r s t o be d u e to π d o n a t i o n f r o m t h e o x y g e n lone pair. T h u s , relative t h e r m o d y n a m i c a n d k i n e t i c considerations appear to b e i m p o r t a n t i n d e t e r m i n a t i o n o f t h e r e a c t i v i t y o f o x o m a n g a n e s e c l u s t e r s . T h i s w o u l d suggest that e n z y m e s that i n c o r p o r a t e a h i g h l y o x y g e n - r i c h l i g a n d e n v i r o n m e n t r e q u i r e a h i g h valent catalytic cycle, whereas those w i t h a n i t r o g e n - r i c h 2

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coordination sphere p r o c e e d t h r o u g h l o w e r valent metal centers. S u c h a p p e a r s to b e t h e case w i t h t h e O E C a n d m a n g a n e s e c a t a l a s e .

Reactivity of [Mn(IV)(salpn)(M -0)] with Acid. A l t h o u g h t h e a d d i t i o n o f a w e a k a c i d w i t h a 2

2

Hydrochloric

noncoordinating a n i o n s u c h as p y r i d i n i u m p e r c h l o r a t e t o [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] l e a d s to t h e s i n g l y p r o t o n a t e d [ M n ( I V ) ( s a l p n ) ^ - 0 ) , ^ - O H ) ] C l 0 (74), s t r o n g acids w i t h c o o r d i n a t i n g anions give drastically different products. T h e m o n o m e r i c Μη(IV) (salpn)Cl can be p r e p a r e d b y the addition of four e q u i v a l e n t s o f H C l to a d i c h l o r o m e t h a n e s o l u t i o n o f [ M n ( I V ) ( s a l p n ) ( μ 0 ) ] . T h i s d e e p g r e e n c o m p l e x , s h o w n i n F i g u r e 6, has trans c h l o r i d e ligation and exhibits a h i g h l y axial E P R s p e c t r u m . T h e details of this conversion have not b e e n t h o r o u g h l y elaborated; h o w e v e r , the m o n o p r o t o n a t e d [ Μ η ( ΐ ν ) ^ 1 ρ η ) ( μ - 0 ) ( μ - Ο Η ) ] a p p e a r s to b e a n i n t e r m e d i a t e o n t h e basis o f l o w - t e m p e r a t u r e s t u d i e s . I f H C l is a d d e d to a d i c h l o r o m e t h a n e s o l u t i o n o f [ M n ( I V ) ( s a l p n ) ( μ - 0 ) ] at - 5 0 ° C , t h e p u r p l e m o n o p r o t o n a t e d d e r i v a t i v e is f o r m e d a n d is s t a b l e f o r s e v e r a l h o u r s . T h i s s o l u t i o n c o n v e r t s d i r e c t l y a n d r a p i d l y to M n ( I V ) ( s a l p n ) C l w h e n t h e s o l u t i o n is w a r m e d to 0 ° C . T h i s d i c h l o r o c o m p l e x is stable i n C H C 1 f o r d a y s at r o o m t e m p e r a t u r e i f p r e c a u t i o n s a r e t a k e n to e x c l u d e c o n t a c t with water. 2

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T h e M n ( I V ) ( s a l p n ) C l is a g o o d o x i d a n t , s h o w i n g a r e d u c t i o n p o t e n t i a l at + 8 9 0 m V v e r s u s S C E . O t h e r h i g h l y o x i d i z e d M n c o m p l e x e s t h a t c o n t a i n c h l o r i n e h a v e b e e n s h o w n to c h l o r i n a t e a l k a n e s , a l k e n e s , a n d 2

Figure 6.

ORTEP diagram of Μη (IV) (salpn) Cl . 2


10.

PECORARO

287


aromatics. F o r this reason, w e d e c i d e d to explore the reactivity of M n ( I V ) ( s a l p n ) C l w i t h a v a r i e t y o f s u b s t r a t e s i n c l u d i n g c y c l o h e x e n e , 1methylcyclohexene, cyclohexane, norbornene, heptyne, and toluene. C y c l o h e x e n e c a n b e c o n v e r t e d to f r a n s - d i c h l o r o c y c l o h e x a n e i n 5 8 % y i e l d after 3 6 h b y M n ( I V ) ( s a l p n ) C l i n C H C 1 . T h e M n c o m p o u n d r e c o v e r e d f r o m t h i s r e a c t i o n is M n ( I I I ) ( s a l p n } C l . N o r b o r n e n e is c o n v e r t e d i n C H C 1 to d i c h l o r o n o r b o r n a n e i n 6 3 % o v e r a l l y i e l d after 2 6 h . T h e p r o d u c t cisitrans r a t i o is 1 : 4 . 7 . M o s t i m p o r t a n t , s k e l e t a l r e a r r a n g e m e n t to g i v e n o r t r i c y c l e n y l c h l o r i d e is n o t o b s e r v e d . B e c a u s e t h e s e c a r b o n m i g r a t i o n s a r e o b s e r v e d w h e n a c a r b o c a t i o n i n t e r m e d i a t e is g e n erated, this eliminates an i o n i c c h l o r i n a t i o n p a t h w a y for the M n ( s a l p n ) b a s e d r e a c t i o n . N e i t h e r t o l u e n e , c y c l o h e x a n e , or h e p t y n e are h a l o g e n a t e d b y M n ( I V ) ( s a l p n ) C l . T h e r e f o r e , c h l o r i n e r a d i c a l ( C l ) is e x c l u d e d as a n i n t e r m e d i a t e b e c a u s e t h i s a g e n t c a n a b s t r a c t h y d r o g e n a n d w i l l c h l o r i n a t e these substrates. A d d i t i o n a l e v i d e n c e s p e a k i n g against a s o l v e n t - d e r i v e d h a l i d e r a d i c a l is t h a t c h l o r i n a t i o n o f o l e f i n s i n C H B r p r o ceeds i n h i g h y i e l d w i t h o u t concomitant b r o m i n a t i o n of substrate. W e f a v o r a r a d i c a l p r o c e s s i n w h i c h t h e m e t a l d i r e c t s t h e final s t e r e o c h e m i s t r y o f t h e p r o d u c t . A p r o p o s e d r e a c t i o n c y c l e is p r o v i d e d as F i g u r e 7. It is p a r t i c u l a r l y n o t e w o r t h y t h a t e a c h o f t h e m o l e c u l e s s p e c i f i e d i n t h i s cycle were isolated and characterized using diffraction techniques. 2

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F i g u r e 7 also s h o w s a n i m p o r t a n t c o m p e t i n g side r e a c t i o n that l o w e r s t h e c h l o r i n a t i o n y i e l d . T h i s is t h e r e a c t i o n o f w a t e r w i t h M n ( I V ) ( s a l p n ) C l giving a Mn(III) product. A l t h o u g h this d e c o m p o s i t i o n of the c o m p l e x is a n a n n o y a n c e i n t h e c h l o r i n a t i o n r e a c t i o n s , it m a y b e o f c o n s i d e r a b l e i n t e r e s t as a p r o c e s s to g e n e r a t e d i o x y g e n f r o m w a t e r . 2

M a t s u s h i t a a n d S h o n o (78, 79) d e m o n s t r a t e d t h a t M n ( I V ) L C l c o m p l e x e s ( w h e r e L is N - a l k y l - 3 - n i t r o s a l i c y l i d e n e i m i n e s ) c a n p r o d u c e o x y g e n f r o m water. T h e reaction of water w i t h these high-valent Schiff-base complexes p r o d u c e d oxygen w i t h a m a x i m u m y i e l d of 0.27 m o l 0 / Mn(IV) complex. Using various N-substituted derivatives, they correlated the reduction potentials of the M n ( I V ) complexes and the h y d r o p h o b i c i t y o f t h e M n ( I V ) c e n t e r to t h e y i e l d o f o x y g e n p r o d u c t i o n (79). M a t s u s h i t a a n d S h o n o also d e m o n s t r a t e d that a m a x i m u m y i e l d is o b t a i n e d at n e u t r a l p H a n d that b y u s i n g O - l a b e l e d H 0 , the o x y g e n p r o d u c e d does c o m e from the water reacting w i t h the M n ( I V ) L C l complexes. T h e y chara c t e r i z e d the products of all of their reactions a n d propose that the r e a c t i o n w i t h w a t e r p r o c e e d s a c c o r d i n g to the f o l l o w i n g e q u a t i o n : 2

2

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l s

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2Mn(IV)L Cl 2

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+ 2 H 0 — 2Mn(II)L + 0 2

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+ 4H

+

+ 4C1"

D e t a i l s o f the w a t e r o x i d a t i o n m e c h a n i s m are u n a v a i l a b l e ; h o w e v e r , it is l i k e l y t h a t o n e c o u l d g e n e r a t e a n i n t e r m e d i a t e H O C 1 t h a t c o u l d go o n , u n d e r a c i d c o n d i t i o n s i n a s e c o n d s t e p , to f o r m 0 . T h i s is r e m i n i s c e n t 2



288


Figure 7.

The proposed chlorination cycle for alkenes by Mn(IV) (salpn) Cl . 2

of the Ru-based water oxidation system of M e y e r , i n w h i c h both 0 w e r e d e t e c t e d (80, 81).

2

and

R e c o g n i z i n g t h e r e l e v a n c e o f b o t h t h e B o u c h e r a n d C o e (72) ([Mn(IV)(Bu-salpn)(M -0)] + H C 1 0 H 0 ) a n d M a t s h u s h i t a et a l . (78, 79) ( M n ( I V ) ( N - P r - 3 - N 0 - s a l ) C l + 2 H 0 0 + 4 H + 4C1") reactions, one can d e d u c e t w o distinct mechanisms for p r o d u c i n g diox y g e n f r o m a [ Μ η ( ΐ ν ) ( μ - 0 ) ] c o r e . I n a d d i t i o n , t h e catalase a c t i v i t y o f [ M n ( I V ) (salpn) ( μ - 0 ) ] d e s c r i b e d b y L a r s o n a n d P e c o r a r o (73) p r o v i d e s a t i d y e x p l a n a t i o n for the alternate catalase a c t i v i t y o b s e r v e d for the O E C . T h e s e t h r e e p r o c e s s e s a r e s h o w n i n F i g u r e 8. 2

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2

2

T h e O E C catalase a c t i v i t y is s u m m a r i z e d i n F i g u r e 8 A , a s s u m i n g a d i m e r of dimers f o r m u l a t i o n for the manganese cluster. Because the c y c l e is b e t w e e n S a n d S , t h e r e l e v a n t e n z y m e o x i d a t i o n l e v e l s a r e M n ( I I I ) M n ( I V ) a n d M n ( I I I ) M n ( I V ) , r e s p e c t i v e l y . T h e catalase c h e m i s t r y is i l l u s t r a t e d at t h e M n ( I V ) d i m e r . T h e first e q u i v a l e n t o f h y d r o g e n p e r o x i d e is o x i d i z e d g i v i n g d i o x y g e n a n d t h e r e d u c e d c l u s t e r . A s u b s e q u e n t m o l e c u l e o f H 0 r e o x i d i z e s t h e c e n t e r to r e g e n e r a t e S . I n t h i s 2

0

3

3

2

2

2


2

10.


PECORARO

'ton

Mrvinj

H2^2 0>

1

9

^

Mni

^

'

H 2^2 0 2

:

289

1^

'—ST""


Mn

Mriî

Mn'

Mn'

2!t) ' 2

Figure 8. The potential relevance of [Mn(IV) (salpn) (μ -0)] catalase ac tivity to the reactions of the oxygen-evolving complex. (A) An explanation for the alternate catalase activity (S S cycle of the oxygen-evolving complex). (B) A mechanism for water oxidation to dioxygen using a com bination of the Boucher and Coe (72) hydrogen peroxide producing reaction and the oxidation of peroxide by [Mn(IV) (salpn) (μ -0)] . (C) A mechanism for water oxidation to dioxygen using the Μη (IV) (salpn) Cl formation re action and the oxidative chemistry of Matsushita et al. (78). The isotopelabeling pattern for the resulting dioxygen differs significantly between mechanisms Β and C. Thefilledoxygens represent 0, and the open oxygens represent 0. 2

0

2

2

2

2

2

18

16

c y c l e d i o x y g e n r e t a i n s t h e l a b e l f o u n d i n p e r o x i d e , as h a s b e e n s h o w n b o t h f o r t h e [ M n ( I V ) (salpn) ( μ - 0 ) ] a n d O E C r e a c t i o n s (47, 72). T h e second process, illustrated i n Figure 8 B , combines the B o u c h e r a n d C o e a c i d i f i c a t i o n c h e m i s t r y (72) w i t h t h e [ M n ( I V ) ( s a l p n ) ^ - 0 ) ] c a t a l a s e c h e m i s t r y (73) t o g e n e r a t e d i o x y g e n . I t r e l i e s on a two-step reaction sequence i n w h i c h a [Μη(ΐν)(μ -0)] reacts w i t h a c i d to generate 2Mn(III) a n d H 0 . A l t h o u g h B o u c h e r a n d C o e did not follow the reaction using labeled [ M n ( I V ) ( B u - s a l p n ) ^ - 0 ) ] , it is l i k e l y t h a t t h i s is s i m p l y t h e b a c k r e a c t i o n o f t h e [ M n ( I V ) (salpn) ( μ 0 ) ] f o r m a t i o n w i t h h y d r o g e n p e r o x i d e (i.e., 2 M n ( I I I ) + H 0 Μ η ( Ι Υ ) ( μ - 0 ) + 2 H ) . I n this case, b o t h oxygens i n t h e r e s u l t a n t h y 2

2

2

2

2

2

2

2

2

2

2

2

2

2

+


2

290


d r o g e n p e r o x i d e w o u l d originate f r o m the same [ Μ η ( ΐ ν ) ( μ - 0 ) ] m o l ecule. Subsequent oxidative chemistry of hydrogen peroxide w i t h a second [ Μ η ( ΐ ν ) ( μ - 0 ) ] w o u l d give 0 + 2Mn(III) along the lines re p o r t e d b y L a r s o n a n d P e c o r a r o (73). F u n c t i o n a l l y , t h i s c o r r e s p o n d s t o c o n v e r s i o n o f O " ( H 0 ) t o d i o x y g e n at t h e e x p e n s e o f f o u r m a n ganese-oxidizing equivalents. F u r t h e r m o r e , i f the initial core was l a b e l e d as [ Μ η ( ΐ ν ) ( μ - 0 ) ] , t h e l i b e r a t e d d i o x y g e n s h o u l d b e e x c l u sively o . 2

2

2

2

2

2

2

1 8

2

1 8

2

2

T h e d i o x y g e n - p r o d u c i n g r e a c t i o n that p r o c e e d s t h r o u g h a m o n o m e r i c M n ( I V ) C l is m e c h a n i s t i c a l l y d i s s i m i l a r to t h e p e r o x i d e p a t h w a y d e s c r i b e d i n t h e p r e c e d i n g p a r a g r a p h a n d s h o u l d l e a d to m i x e d - l a b e l e d 0 . I n t h e first s t e p , a c i d a n d h a l i d e a t t a c k t h e [ Μ η ( ΐ ν ) ( μ - 0 ) ] c o r e as we described i n preceding paragraphs; however, rather than liberating H 0 i n a r e d o x r e a c t i o n , t h e M n ( I V ) C l i n t e r m e d i a t e is f o r m e d a n d w a t e r is r e l e a s e d . T h i s H 0 c a n t h e n b a c k r e a c t w i t h t h e M n ( I V ) C l t o g i v e 0 a c c o r d i n g to t h e c h e m i s t r y o f M a t s u s h i t a et a l . (78). A n o x y g e n label w o u l d scramble i n this reaction because all of the oxygen atoms are r e l e a s e d p r i o r to r e o x i d a t i o n . A n o t h e r i m p o r t a n t d i s t i n c t i o n b e t w e e n t h e B o u c h e r - C o e a n d M a t s u s h i t a i n t e r m e d i a t e s is t h a t i n t h e first case t w o s e q u e n t i a l t w o - e l e c t r o n o x i d a t i o n s at m a n g a n e s e d i m e r s l e a d to product, whereas i n the second, acid-base chemistry precedes oxidation t h a t o c c u r s e x c l u s i v e l y at m o n o n u c l e a r c e n t e r s .


2

2

2

2

2

2

2

2

2

2

Dinuclear Manganese Complexes as Models for the Man ganese Catalase. A s d i s c u s s e d p r e v i o u s l y , t h e m a n g a n e s e catalase has a d i n u c l e a r a c t i v e site that is t h o u g h t to f u n c t i o n b y c y c l i n g r e d o x states between M n ( I I ) and Mn(III) . A l t h o u g h the [Mn(IV)(salpn)(μ -0)] c h e m i s t r y n i c e l y e x p l a i n s t h e a l t e r n a t e catalase r e a c t i o n s o f t h e O E C , t h i s s y s t e m is a n i n a p p r o p r i a t e m o d e l f o r t h e M n catalase b e c a u s e t h e r e d o x c y c l e i n t h a t e n z y m e is l o w e r a n d t h e c o r e s t r u c t u r e is b e l i e v e d to b e d r a m a t i c a l l y d i f f e r e n t . I n f a c t , a [ Μ η ( Ι Ι Ι / ΐ ν ) ( μ - 0 ) ] s u p e r o x i d i z e d state o f t h e M n catalase has b e e n i d e n t i f i e d a n d s h o w n t o b e i n a c t i v e . 2

2

2

2

2

2

W e o b s e r v e d t h a t a s l i g h t m o d i f i c a t i o n o f t h e s a l p n l i g a n d l e d to d r a m a t i c a l l y d i f f e r e n t c o o r d i n a t i o n c h e m i s t r y t h a t has p r o v i d e d s i g n i f i c a n t i n s i g h t i n t o t h e m a n g a n e s e catalase r e a c t i o n s . S u b s t i t u t i o n o f a h y d r o x i d e g r o u p at t h e 2 p o s i t i o n o f t h e p r o p a n e b a c k b o n e l e a d s to t h e pentadentate ligand 2 - O H - s a l p n . T h r e e distinct dinuclear structure t y p e s , i l l u s t r a t e d i n F i g u r e 9, h a v e b e e n d e f i n e d f o r m a n g a n e s e w i t h t h i s l i g a n d . T h e first c o n t a i n s a d i - μ - ο χ ο b r i d g e d c o r e t h a t is a n a l o g o u s to [ M n ( I I I / I V ) ( s a l p n ) ^ - 0 ) ] ~ . T h e s e c o n d a n d t h i r d categories have the 2 - h y d r o x y l g r o u p b o u n d as a n a l k o x i d e . G r o u p t w o c o r r e s p o n d s t o s y m m e t r i c s t r u c t u r e s , so d e s i g n a t e d b e c a u s e t h e m o l e c u l e s c o n t a i n t w o , equivalent b r i d g i n g alkoxide donors that are part o f the l i g a n d . T h e t h i r d category contains a s y m m e t r i c c o m p l e x e s that have o n l y one o f the 2

2


PECORARO


291


10.

\

{Mn(m)(2-OHSALPN)(CH OH] 3

2

Figure 9. Three distinct structural types are available for Mn complexes of 2-OH-salpn. (A) A di-μ-οχο bridged core structure represented by [Mn(IIIJV) (2-OH-salpn) (μ -0)] ~. (Β) Symmetric dialkoxide compounds represented by the [Mn(III)(2-OH-salpn)] . (C) Asymmetric dialkoxide compounds represented by [Mn(III) (2-OH-salpn) (CH OH)]. 2

2

2

2

2

3


292


alkoxide oxygen atoms p a r t i c i p a t i n g i n a b r i d g e b e t w e e n the manganese i o n s (82, 83). A l t h o u g h c a t e g o r y 1 a n d 2 c o m p l e x e s h a v e a l l m a n g a n e s e c o o r d i n a t i o n sites filled b y t h e l i g a n d o r μ - 0 b r i d g e s , t h e s i x t h c o o r d i n a t i o n site o f o n e o f t h e m a n g a n e s e i o n s i n t h e c a t e g o r y 3 s t r u c t u r e s is filled b y s o l v e n t . T h e s e t h r e e s t r u c t u r e t y p e s s h o w a s u b s t a n t i a l r a n g e of M n - M n separations. T h e [Mn(III/IV)(2-OH-salpn) ( μ - 0 ) ] ~ structure is e x p e c t e d t o h a v e a M n - M n s e p a r a t i o n o n t h e o r d e r o f 2.7 À . I n c o n trast, t h e M n ( I I I / I V ) d i s t a n c e i n t h e a s y m m e t r i c c o m p l e x [ M n ( I I I / I V ) ( 2 O H - s a l p n ) T H F ] is 3 . 6 1 À (84). R e d u c i n g t h i s c o m p l e x b y o n e e l e c t r o n leads to the a s y m m e t r i c [ M n ( I I I ) ( 2 - O H - s a l p n ) C H O H ] ( M n - M n 3.83 À ) . T h i s is o v e r 0 . 5 À l o n g e r t h a n t h e s y m m e t r i c d i m e r [ M n ( I I I ) ( 2 - O H s a l p n ) ^ ( M n - M n 3 . 2 3 À ) (85). S t e p w i s e r e d u c t i o n o f t h e s y m m e t r i c M n ( I I I ) d i m e r r e s u l t s i n t h e l e n g t h e n i n g o f t h e M n - M n d i s t a n c e , first to [ M n ( I I , I I I ) ( 2 - O H - s a l p n ) ] - ( M n - M n 3 . 3 7 À ) a n d t h e n a s m a l l d e c r e a s e to [ M n ( I I ) ( 2 - O H - s a l p n ) ] - ( M n - M n 3.33 À ) . 2

2

2

2

+

2


2

2

3

2

2

2

T h e s y m m e t r i c [ M n ( I I ) ( 2 - O H - s a l p n ) ] ~ is a i r - s e n s i t i v e a n d w i l l c o n v e r t to [ M n ( I I I ) ( 2 - O H - s a l p n ) ] in acetonitrile and [Mn(III) (2-OHs a l p n ) C H O H ] in methanol. A wide variety of p h e n y l ring derivatives c a n b e p r e p a r e d t h a t a l l o w f o r t h e fine t u n i n g o f t h e r e d o x p o t e n t i a l s o f t h e s e m a n g a n e s e c o m p l e x e s . I f t h e [ M n ( I I ) ( 2 - O H - ( 3 , 5 - C l - s a l ) p n ) ] ~ is a i r - o x i d i z e d at - 4 0 ° C i n e t h a n o l , t h e i n t e r m e d i a t e [ M n ( I I , I I I ) ( 2 - O H (3,5-Cl-sal)pn)] ~ can be isolated i n high yield. Alternatively, bleeding a s m a l l a m o u n t o f 0 i n t o a r e a c t i o n v e s s e l at r o o m t e m p e r a t u r e w i l l provide [Mn(II,III)(2-OH-salpn)] -, but i n l o w y i e l d . T h e [Mn(III)(2O H - s a l p n ) ] can be converted into the asymmetric complex, [Mn(III) (2O H - s a l p n ) C H O H ] , by dissolving i n methanol. T h e desolvation of [ M n ( I I I ) ( 2 - O H - s a l p n ) C H O H ] to give the s y m m e t r i c d i m e r c a n b e 2

2

2

2

2

3

2

2

2

2

2

2

2

2

3

2

2

3

achieved by redissolving in acetonitrile. T h e reaction of hydrogen peroxide w i t h [Mn(II)(2-OH-salpn)] ~ or [ M n ( I I I ) ( 2 - O H - s a l p n ) ] i n a c e t o n i t r i l e causes the e v o l u t i o n of d i o x y g e n , a p p a r e n t l y c y c l i n g b e t w e e n t h e t w o c o m p l e x e s (85). O x y g e n e v o l u t i o n e x p e r i m e n t s i n d i c a t e that these c o m p l e x e s can c o m p l e t e the catalase r e a c t i o n f o r at least 1 0 0 0 t u r n o v e r s w i t h o u t s i g n i f i c a n t d e c o m p o s i t i o n o f the catalyst. Isolation o f d i o x y g e n after the a d d i t i o n of H 0 yields exclusively 0 . If H 0 and H 0 are a d d e d to either manganese complex, 0 and 0 are r e c o v e r e d but no 0 is d e t e c t e d . T h i s isotope-labeling follows the isotopic 0 c o m p o s i t i o n s h o w n for the L . plantarum c a t a l a s e . T h e r e a c t i o n o f h y d r o g e n p e r o x i d e w i t h a 5 0 : 5 0 mixture of [Mn(II)(2-OH-salpn)] - and [Mn(II)(2-OH-(5-N0 -sal)pn)] g i v e s mass p e a k s f r o m t h e r e c o v e r e d m a t e r i a l o n l y f o r [ M n ( I I I ) ( 2 - O H salpn)] and [Mn(III)(2-OH-(5-N0 -sal)pn)] , and no [ M n ( I I I ) ( 2 - O H ( s a l p n ) ( 2 - O H - ( 5 - N 0 - s a l ) p n ) j . T h i s provides strong e v i d e n c e that the d i m e r s are not dissociating into m o n o m e r s d u r i n g the catalytic process. T h e U V - v i s i b l e ( U V - v i s ) s p e c t r a o f the catalase r e a c t i o n u p o n a d d i t i o n 2

2

2

1 8

2

1 8

1 8

2

2

2

1 6

1 8

2

1 6

2

2

2

1

2

6

1

8

2

2

2

2

2

2

2

2

2


2

2

2

10.

PECORARO

293


o f 1 e q H 0 t o [ M n ( I I I ) ( 2 - O H - s a l p n ) ] a r e s h o w n as F i g u r e 1 0 B . F i g u r e s 1 OA a n d 1 0 C show the spectra for [ M n ( I I ) ( 2 - O H - s a l p n ) ] - a n d [Mn(III)(2-OH-salpn)] , respectively. T h e isosbestic point demonstrates a c l e a n c o n v e r s i o n f r o m the [ M n ( I I I ) ( 2 - O H - s a l p n ) ] to the [Mn(II)(2O H - s a l p n ) ] ~ . T h i s conversion shows no evidence for an i n t e r m e d i a t e . 2

2

2

2

2

2

2

2

2

T h e initial rate of reaction of these complexes w i t h H 0 was c a l c u l a t e d u s i n g a m o d i f i e d C l a r k - t y p e 0 e l e c t r o d e . T a b l e II c o m p a r e s the initial rate of different derivatives of [Mn(II)(2-OH-salpn)] ~ a n d [ M n ( I I I ) ( 2 - O H - s a l p n ) ] . C l e a r l y , t h e o b s e r v e d rates f o r a l l o f t h e d e r i v atives w e r e v e r y similar. W e d e c i d e d to u n d e r t a k e a d e t a i l e d k i n e t i c s t u d y o f t h e [ M n ( I I I ) ( 2 - O H - ( 5 - C l - s a l ) p n ) ] b e c a u s e t h i s m o l e c u l e has t h e greatest s o l u b i l i t y i n a c e t o n i t r i l e . F i g u r e 1 1 A s h o w s that t h i s c o m p l e x exhibits saturation k i n e t i c s w i t h respect to h y d r o g e n p e r o x i d e , w h e r e a s F i g u r e 1 2 d e m o n s t r a t e s t h a t at s a t u r a t i n g h y d r o g e n p e r o x i d e c o n c e n t r a t i o n s t h e r e a c t i o n is first o r d e r i n [ M n ( I I I ) ( 2 - O H - ( 5 - C l - s a l ) p n ) ] . T h e s a t u r a t i o n - t y p e k i n e t i c s suggest t h a t t h e r e is a r e v e r s i b l y f o r m e d " p e r o x y [ M n ( 2 - O H - ( 5 - C l - s a l ) p n ) ] " intermediate i n the t u r n o v e r l i m i t i n g step; 2

2

2

2

2

2


2

2

2

2.0 H «

2.0 π .

2.0 H

Wavelength (nm)

Figure 10. (A) UV-vis spectrum ofNa [Mn(II) (2-OH-(3,5-Cl-sal)pn)] . (B) Conversion of [Mn(III) (2-OH-(3,5-Cl-sal)pn)] to [Mn(II)(2-OH-(3,5-Clsal)pn)] ~ by addition of 1 equiv ofH 0 . (C) UV-vis spectrum of[Mn(III) (2OH-(3,5-Cl-sal)pn)] . 2

2

2

2

2

2

2


294


Table II.

Initial Rate Data for M n Catalase and M n Models Rate (mol H 0 /mol catalyst s' )

Catalyst

2

L. plantarum Mn catalase Na [Mn(II)(2-OH-(3,5-Cl-sal)pn)] [Mn(ffl)(2-OH-(3,5-Cl-sal)pn)] [Mn(III)(2-OH-(5-Cl-sal)pn) ] Na [Mn(II)(2-OH-(5-Cl-sal)pn)] [Mn(III)(2-OH-salpn)] [Mn(ffl)(2-OH-(5-N0 -sal)pn)] Mn(II)(C10 ) -6H 0 2

2 11.0 10.0 12.4 13.1 13.0 11.2 6.25

2

2

2

2

2

2

2


4

2

2

2

1

2

Χ 10 ±0.9 ± 0.8 ±0.5 ±0.4 ± 1.1 ± 0.5 ± 1.5 X 1(T 5

3

h o w e v e r , the U V - v i s spectra show no evidence of this intermediate. A d o u b l e r e c i p r o c a l p l o t o f t h e d a t a i n F i g u r e 11 A , s h o w n as F i g u r e 1 1 B , d e m o n s t r a t e s a l i n e a r r e l a t i o n s h i p f r o m w h i c h fc a n d K m a y b e c a l c u l a t e d . T h e K ( H 0 ) = 3 7 ± 1 0 m M a n d fc = 1 3 ± 3 m o l H 0 c o n s u m e d / s . I n c o m p a r i s o n , t h e M n c a t a l a s e f r o m L. plantarum has a K ( H 0 ) = 2 0 0 m M a n d fc = 2 Χ 1 0 m o l H 0 c o n s u m e d / s , w h e r e a s M n ( I I ) ( C l 0 ) has fc = 6 . 3 Χ 1 0 " m o l H 0 c o n s u m e d / s . R e l a t i v e v a l u e s f o r k /K for the M n catalase a n d [ M n ( I I I ) ( 2 - O H - ( 5 - C l - s a l ) p n ) ] a r e 1 Χ 1 0 a n d 3.5 Χ 1 0 , r e s p e c t i v e l y , i n d i c a t i n g t h a t t h e e n z y m e is ~ 3 0 0 0 times m o r e efficient t h a n o u r synthetic catalyst. m

cat

m

m

2

2

2

cat

2

5

cat

4

cat

2

2

3

cat

2

2

2

2

2

m

2

6

2

T h e L. plantarum catalase w a s d e s c r i b e d i n i t i a l l y as a z i d e i n s e n s i t i v e b e c a u s e t h e r e a c t i o n is n o t i n h i b i t e d at a z i d e c o n c e n t r a t i o n s t h a t w o u l d e a s i l y d e s t r o y h e m e c a t a l a s e a c t i v i t y . H o w e v e r , P e n n e r - H a h n (22) has s h o w n that this e n z y m e can b e i n h i b i t e d b y azide i f the concentrations a r e s u f f i c i e n t l y h i g h . T h e K = 8 0 m M ( p H 7) a n d is p H d e p e n d e n t . T h e [ M n ( I I I ) ( 2 - O H - ( 5 - C l - s a l ) p n ) ] is n o t i n h i b i t e d i n a c e t o n i t r i l e to s a t u r a t i n g concentrations of azide ( « 5 0 m M ) . {

2

H y d r o x y l a m i n e is a n o t h e r p e r o x i d e a n a l o g u e t h a t has b e e n u s e d as a p r o b e o f e n z y m e a c t i v i t y . I f t h e M n c a t a l a s e is e x p o s e d t o N H O H i n t h e a b s e n c e o f h y d r o g e n p e r o x i d e , t h e e n z y m e is r e d u c e d t o t h e M n ( I I ) f o r m . H o w e v e r , i f h y d r o g e n p e r o x i d e is p r e s e n t , t h e e n z y m e is t r a p p e d in a catalytically inactive, superoxidized Mn(III,IV) form. Treatment w i t h N H O H i n the absence of h y d r o g e n peroxide w i l l regenerate the r e d u c e d f o r m a n d a c t i v i t y . T h e [ M n ( I I ) ( 2 - O H - s a l p n ) ] ~ s y s t e m c a n also b e d r i v e n t o a c a t a l y t i c a l l y i n a c t i v e M n ( I I I , I V ) f o r m t h a t has a n E P R s p e c t r u m that is s t r i k i n g l y s i m i l a r t o t h e s p e c t r a o f b o t h t h e s u p e r o x i d i z e d M n catalase a n d [ M n ( I I I , I V ) (salpn) ( μ - 0 ) ] ~ . It is i m p o r t a n t to n o t e t h a t t h e E P R s p e c t r u m o f this c o m p l e x is d i f f e r e n t f r o m t h a t o f t h e a s y m m e t r i c c o m p l e x [ M n ( I I I , I V ) ( 2 - O H - s a l p n ) T H F ] . T h e four E P R spectra are c o m p a r e d i n F i g u r e 1 3 . A s is t h e case w i t h t h e e n z y m e , t h e [ M n ( I I I , I V ) ( 2 - O H - ( s a l p n ) ] (this m o l e c u l e is e i t h e r [ M n ( I I I , I V ) ( 2 - O H 2

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Reactivity and Mechanism of Manganese Enzymes - Advances in

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