Ion Pumps as Energy Transducers - ACS Symposium Series (ACS

14

Ion Pumps as Energy Transducers ERICH HEINZ

Downloaded by RUTGERS UNIV on May 30, 2018 | https://pubs.acs.org Publication Date: January 18, 1983 | doi: 10.1021/bk-1983-0207.ch014

Cornell University Medical College, New York, NY 10021

Ionmotive forces, i.e. the electrochemical potential differences produced by ion pumps may serve to transmit energy released by exergonic reactions to drive endergonic processes, such as synthesis of energy-rich compounds o r uphill transport of organic solutes. A typical example is the "chemiosmotic coupling" by a proton pump in oxidative and photosynthetic phosphorylation. The effectiveness of this coupling depends on its (energetic) efficiency as well as on its (kinetic) expeditiousness. The efficiency is mainly limited by inner and outer leakages to the transported ion species, the expeditiousness mainly by the delay in generating the protonmotive force. Whereas little is known about the various leakages to assess the efficiency, some preliminary information concerning the expeditiousness may be considered here. The generation of an ionmotive force, to the extent that it depends on ion gradients is a rather slow process. Electrogenic pumps, however, can more rapidly raise the electrical potential, p r i o r to appreciable ion translocation by loading the static membrane capacity. Such rapid electric effects may significantly increase the expeditiousness of the coupling but only under the condition that the pump works at constant driving force. On the other hand, i f the pump works at constant rate the r i s e of the protonmotive force is largely determined by the formation of ion gradients which r i s e more slowly and are less sensitive towards variations in energy input. Using a recently developed optical method of rapidly monitoring the P D , it could be shown that the light-driven proton pump of halobacterium halobium has the high expeditiousness predicted for an electrogenic pump working at constant driving force.

0097-6156/83/0207-0323$06.00/0 © 1983 American Chemical Society

Blanch et al.; Foundations of Biochemical Engineering ACS Symposium Series; American Chemical Society: Washington, DC, 1983.

324

BIOCHEMICAL

In o r d e r cesses

that e n e r g y be t r a n s f e r r e d

these have

to b e

two c h e m i c a l r e a c t i o n s

coupled.

ENGINEERING

b e t w e e n two d i f f e r e n t

Homogenous

c o u p l i n g , i . e.

w i t h i n the s a m e c o m p a r t m e n t i s

pro

between

usually

b a s e d on the p r i n c i p a l of the " c o m m o n i n t e r m e d i a t e " a s i s i l l u s t r a t e d b y the " c l a s s i c a l " c o u p l i n g b e t w e e n the d e h y d r o g e n a t i o n phosphate

diphosphoglycerate l i n k s t h e s e two

was

identified as

reactions.

A n analagous,


or a mobile

supposedly

gate.

More

formed

b y the two

c o m p l i c a t e d and l e s s

the c o r r e s p o n d i n g p r i n c i p l e of h e t e r o g e n o u s c h e m i c a l and an o s m o t i c p r o c e s s has been discussed

the r e v e r s e

processes,

namely through

solutes

with a

well understood

until

for m i t o c h o n d r i a l and b a c t e r i a l " o x i d a t i v e r i c h phosphate

phosphory should

the

be

In the e v o l v i n g

" c h e m i o s m o t i c h y p o t h e s i s " o f o x i d a t i v e p h o s p h o r y l a t i o n two coupling o c c u r in s e r i e s :

first

s t e p s of

one l i n k s a n o x i

d a t i o n - r e d u c t i o n r e a c t i o n to the a c t i v e e x t r u s i o n o f p r o t o n s , generating an e l e c t r o c h e m i c a l potential difference ("protonmotive

force");

phosphate b y means There is

of

to the s y n t h e s i s of

r e e n t r y of

energy-rich

o f the " p r o t o n t r a n s l o c a t i n g A T P a s e "

good e v i d e n c e

that the s a m e s e q u e n c e

thereby

H-ions

the s e c o n d one l i n k s the p a s s i v e

d r i v e n by this force,

(Fig.

s t e p s o c c u r s i n the o x i d a t i v e p h o s p h o r y l a t i o n o f a e r o b i c

s y s t e m s have

Various plausible

bacteria

models

of

s i n c e been d e v i s e d and c r i t i c a l l y d i s c u s s e d

evidence

it is

so far

s u r p r i s i n g that t h i s c o u p l i n g , cal

well

enough supported b y

to j u s t i f y the wide b e l i e f i n i t s v a l i d i t y .

such

In o t h e r w o r d s ,

two " c h e m i o s m o t i c e n e r g y a mere

advantages as c o m p a r e d

experimental

not i n v o l v e

b u t r a t h e r two h e t e r o g e n o u s energy is transferred

r e a c t i o n to a n o t h e r one i n o s m o t i c f o r m ,

k i n d of c o u p l i n g i s

8, 9 ). proven

w h i c h u l t i m a t e l y l i n k s two t r u l y c h e m i

common chemical intermediate, steps i n s e r i e s .

(3) 5 ),

It i s n o n e t h e l e s s

r e a c t i o n s , d o e s i n c o n t r a s t to the c l a s s i c a l m o d e l s

chemical

(4,

(7,

T h o u g h the p o s t u l a t e d " c h e m i o s m o t i c " c o u p l i n g h a s not b e e n y e t b e y o n d doubt,

1).

of s i m i l a r c o u p l i n g

a s w e l l a s i n the l i g h t - d r i v e n p h o s p h o r y l a t i o n o f c h l o r o p l a s t s and h a l o p h i l i c b a c t e r i a ( 6 ) .

to

M i t c h e l l ( 2 ) postulated

e n e r g i z e d b y the d o w n h i l l m o v e m e n t o f Η - i o n s .

protons,

as

T h i s coupling had

of t r a n s f e r r i n g c h e m i c a l e n e r g y

osmotic process,

l a t i o n " w h e r e the s y n t h e s i s of e n e r g y

heterogenous

is a

in p r i m a r y active transport, ( 1 ).

a

carrier

coupling,between

in m o r e detail elsewhere

mainly been considered a means drive an endergonic

which

though s o m e w h a t d i f f e r e n t

in (secondary active) ion-linked cotransport,

ternary complex

triose-

for

the " c o m m o n i n t e r m e d i a t e " that

p r i n c i p l e u n d e r l i e s the c o u p l i n g b e t w e e n two o s m o t i c e. g.

of

a n d the f o r m a t i o n of a d e n o n i s i n e t r i p h o s p h a t e ,

transductions". vestige

Whether

o r whether

from

one

with the h e l p of this p e c u l i a r

it has c e r t a i n

to o t h e r k i n d s of c o u p l i n g ,

b e i n g m o r e e f f e c t i v e , is h a r d to t e l l .

a

coupling

for instance,

by

L e t u s s e e how a n d u n d e r w h i c h

conditions a g r e a t e r effectiveness m i g h t c o m e about.



14.

HEINZ

Ion

325

Pumps

Figure 1. Chemiosmotic coupling in oxidative phosphorylation. The model is drawn according to Mitchell's hypothesis for mitochondria, but might also apply to other systems of phosphorylation. A hydrogen ion pump transports the H-ions from left to right, energized by an oxidationreduction reaction, a light-driven photochemical mechanism, or any other appropriate energy source. This pumping is tantamount to pumping Ο H-ions in the opposite direction. A membranebound enzyme, the proton translocating ATPase catalyzes the interconversion of A DP and Pi and ATP by a heterophasic (or vectorial) reaction, i.e., a reaction in which the Ο H-ions, released by the synthesis or incorporated by the hydrolysis of ATP, can interact with the en zyme only from the phase contralateral to that from which the other reactants interact. In this way, the direction of the chemical reaction de pends on the spatial direction of the H(or OH-) gradient.


326

BIOCHEMICAL

T h e o v e r a l l effectiveness

ENGINEERING

of a n y c o u p l i n g m e c h a n i s m m a y b e

a s s e s s e d i n t e r m s o f i t s ( e n e r g e t i c ) e f f i c i e n c y a s w e l l a s i n t e r m s of i t s (kinetic) e x p e d i t i o u s n e s s .

T h e e n e r g e t i c e f f i c i e n c y i s l i m i t e d b y the

e x t e n t of u n a v o i d a b l e l e a k a g e s ,

w h i c h a r e p a r t l y due to p a s s i v e

permea-

b i l i t y of the m e m b r a n e b a r r i e r to the t r a n s p o r t e d s o l u t e (outer l e a k a g e ) a n d p a r t l y r e s i d e i n the t r a n s p o r t m e c h a n i s m i t s e l f ( i n n e r l e a k a g e ) i n that " s l i p p i n g " a n d " b a c k c y c l i n g " m a y p e r m i t the e s c a p e of u n u t i l i z e d i n p u t e n e r g y (1 ).

T o o l i t t l e i s k n o w n about the e x t e n t of l e a k a g e s w i t h -

i n the c h e m i o s m o t i c m e c h a n i s m o f p h o s p h o r y l a t i o n to c o m p a r e i t s e n e r g e t i c e f f i c i e n c y w i t h that o f i t s c h e m i c a l c o u n t e r p a r t .


N o t m u c h m o r e i s known about the e x p e d i t i o u s n e s s w h i c h , i n t h i s c o n t e x t , m a y be d e f i n e d a s the r a t e at w h i c h the p r o t o n m o t i v e f o r c e r i s e s a f t e r the o n s e t of the p u m p , but s o m e g e n e r a l p r e d i c t i o n s c a n be m a d e .

T h e e x p e d i t i o u s n e s s i s the h i g h e r the f a s t e r the p u m p i n g

r a t e of p r o t o n s i s r e l a t i v e to the c o n d u c t a n c e o f p a s s i v e i o n s , tend to shunt the e l e c t r o g e n i c p u m p effect, electroneutrality. force,

which

i n o t h e r w o r d s , to m a i n t a i n

W e s e e that the two c o m p o n e n t s of the p r o t o n m o t i v e

the e l e c t r i c a l a n d the c h e m i c a l P D , a r e c o n t r o l l e d b y d i f f e r e n t

f a c t o r s a n d m a y t h e r e f o r e d e v e l o p at d i f f e r e n t r a t e s , a s i s i l l u s t r a t e d b y the f o l l o w i n g two e x t r e m e c o n d i t i o n s : If the p a s s i v e c o n d u c t a n c e i s e x t r e m e l y l o w ,

the e l e c t r i c a l

p o t e n t i a l w i l l b e g e n e r a t e d p u r e l y e l e c t r o g e n i c a l l y , i . e. a p p r e c i a b l e net m o v e m e n t of p a s s i v e i o n s . the m e m b r a n e i s r a t h e r l o w ,

^F/cm , 2

without

A s the e l e c t r i c c a p a c i t y of

e l e c t r o g e n i c e x p u l s i o n of 3 0 0 -

400 p r o t o n s p e r c e l l , p a s s i v e s h u n t i n g effect b e i n g n e g l i g i b l e at t h i s stage,

s h o u l d s u f f i c e to r a i s e the e l e c t r i c a l P D b y 1 m v o l t . O n the o t h e r h a n d , i f the p a s s i v e c o n d u c t a n c e i s v e r y h i g h , i t

w i l l k e e p e l e c t r i c a l e f f e c t s s m a l l but a l l o w i n s t e a d a n e q u i v a l e n t r i s e o f the c h e m i c a l P D of p r o t o n s .

T h i s r i s e w i l l d e p e n d o n the b u f f e r

c a p a c i t y o f the m e d i u m , w h i c h i s n o r m a l l y m u c h h i g h e r than the e l e c t r i c a l capacity.

F r o m i t s v a l u e i n the m i t o c h o n d r i a we w o u l d e s t i -

m a t e that about a m i l l i o n p r o t o n s h a v e to be e x p e l l e d p e r 1 m v o l t r i s e of the c h e m i c a l P D , a n d h e n c e o f the p r o t o n m o t i v e f o r c e .

Obviously,

the r i s e i n c h e m i c a l P D i n t h i s c a s e i s t h o u s a n d o r m o r e t i m e s s l o w e r than the e l e c t r o g e n i c one i n the f o r m e r c a s e .

A t the r a t e a s s u m e d

for

the m i t o c h o n d r i a l p r o t o n p u m p , we would p r e d i c t that the c r i t i c a l protonmotive force,

i . e . about 200 m v o l t s , c o u l d be r e a c h e d w i t h i n a

few h u n d r e d m i l l i s e c o n d s i n the f i r s t c a s e , but would take a few h u n d r e d s e c o n d s i n the l a s t c a s e .

It f o l l o w s that the e x p e d i t i o u s n e s s of

the c o u p l i n g , i . e. the s p e e d at w h i c h t h i s c r i t i c a l p r o t o n m o t i v e f o r c e i s r e a c h e d a f t e r the i n i t i a t i o n of the p u m p d e p e n d s on the r e l a t i v e c o n t r i b u t i o n of the i n i t i a l p u m p i n g r a t e r e l a t i v e to the s h u n t i n g p a t h w a y s for passive i o n s .


14.

Ion

HEINZ

327

Pumps

T o a p p r o a c h t h i s p r o b l e m q u a n t i t a t i v e l y , we m a y f o r s i m p l i c i t y r e a s o n s f o l l o w the c o n v e n t i o n a l p r o c e d u r e u s e d b y e l e c t r o p h y s i o l o g i s t s i n d e a l i n g with e l e c t r o g e n i c p u m p s in t h e i r b i o l o g i c a l s y s t e m s , e t c .

In

the e q u i v a l e n t c i r c u i t s u s e d b y t h e m to v i s u a l i z e the e l e c t r i c p h e n o m e n a of b i o l o g i c a l s y s t e m s , an electrogenic ion p u m p i s u s u a l l y represented e i t h e r as a constant voltage s o u r c e o r as a constant c u r r e n t s o u r c e , b a s e d o n the s i m p l i f y i n g a s s u m p t i o n that d u r i n g the e x p e r i m e n t a l o b s e r v a t i o n e i t h e r the d r i v i n g f o r c e o f the p u m p remains

fairly constant.

o r the p u m p i n g r a t e

T h e d i f f e r e n c e b e t w e e n t h e s e two

a l t e r n a t i v e s c o r r e s p o n d s w i t h s o m e c r u d e a p p r o x i m a t i o n to the a b o v e d i s c u s s e d d i f f e r e n c e b e t w e e n two k i n d s o f c o u p l i n g m o d e l s ; the c o n s t a n t


v o l t a g e s o u r c e n e t w o r k m o s t c l o s e l y w o u l d a c c o u n t f o r the e x p e d i t i o u s n e s s , but l e s s s t a b l e c o u p l i n g m o d e l , w h e r e a s the c o n s t a n t c u r r e n t s o u r c e w o u l d d o s o f o r the m o r e s l u g g i s h b u t mo r e s t a b l e m o d e l . most cases,

In

i t i s d i f f i c u l t t o t e l l a p r i o r i w h i c h o f the two i s c o r r e c t a s

f o r e i t h e r one s o m e p l a u s i b l e e x p l a n a t i o n c a n b e g i v e n .

As a conse-

q u e n c e , the t r e a t m e n t b y e l e c t r o p h y s i o l o g i s t s i n t U s r e s p e c t i s n o t u n i f o r m , b u t a s l o n g a s the s t e a d y s t a t e i s c o n c e r n e d , e i t h e r one m a y give reasonable a n s w e r s . becomes,

T h e d i f f e r e n c e b e t w e e n the two m o d e l s

however, c r u c i a l i n transient states, for instance, i m m e d i -

a t e l y a f t e r the p u m p h a s b e e n t u r n e d o n o r

off.

M i t c h e l l ( 8 ) was p r o b a b l y the f i r s t t o i n v e s t i g a t e t h e s e

questions»

a n d b y r i g o r o u s c a l c u l a t i o n s , b a s e d on the a s s u m p t i o n o f c o n s t a n t p u m p i n g r a t e , h e p r e d i c t e d that the e l e c t r i c a l P D r i s e s v e r y f a s t but i n s i g n i f i c a n t l y h i g h w h i l e the f u l l d e v e l o p m e n t o f the p r o t o n m o t i v e f o r c e h a s to a w a i t the g e n e r a t i o n o f a s u f f i c i e n t l y h i g h c o n c e n t r a t i o n g r a d i e n t w h i c h w o u l d t a k e m o r e t h a n 30 s e c o n d s . f o r the a s s u m p t i o n that the d r i v i n g f o r c e ,

The same approach,

except

r a t h e r t h a n the r a t e o f the

p u m p r e m a i n s c o n s t a n t , g a v e a g r e a t l y d i f f e r e n t r e s u l t (10, 11 ). D e p e n d i n g o n the p u m p i n g r a t e r e l a t i v e to the p a s s i v e i o n m o b i l i t i e s , t h e i n i t i a l s u r g e o f the e l e c t r i c a l p o t e n t i a l m a y i n d e e d b e h i g h e n o u g h to a n t i c i p a t e m o s t o f the final p r o t o n m o t i v e f o r c e a t s t a t i c h e a d ( F i g . 2).

O b v i o u s l y , a c o n s t a n t p u m p i n g r a t e i m p l i e s that the d r i v i n g

f o r c e i s i n i t i a l l y l o w but r i s e s a c c o r d i n g l y a s the o p p o s i n g p r o t o n m o t i v e force grows.

B y contrast, at constant d r i v i n g force,

the p u m p i n g r a t e

s h o u l d i n i t i a l l y b e v e r y h i g h a n d l a t e r d e c l i n e a c c o r d i n g l y a s the opposing protonmotive force r i s e s .

A p r i o r i , both models a r e i n s o m e

w a y p l a u s i b l e a n d o n l y the e x p e r i m e n t c a n d e c i d e w h i c h one i s the v a l i d one. So f a r ,

e x p e r i m e n t a l t e s t s a r e s c a r c e , p a r t l y due t o the

d i f f i c u l t i e s to m e a s u r e r a p i d l y e n o u g h c h a n g e s i n e l e c t r i c a l m e m b r a n e p o t e n t i a l (or p r o t o n m o t i v e f o r c e ) w i t h s m a l l c o m p a r t m e n t s s u c h a s mitochondria and b a c t e r i a .

T h e usually applied chemical probes

p o t e n t i a l c h a n g e s a r e e i t h e r too s l o w o r too n o i s y .

for

Recently, however,


328

BIOCHEMICAL


H + -pump

M

II

on

,,1

II

ENGINEERING

,,ιι

Η -pump o f f

Figure 2. The calculated response of the protonmotive force, and its electrical and chemical components, to the sudden initiation or stopping of an electrogenic proton pump. Ordinate: protonmotive force (X ) in relative units. Abscissa: time. Solid line, electric PD; dashed line, chemical PD of H*; dotted line, X . Top: constant rate of pumping (constant current source). Bottom: constant driving force of the pump (constant voltage source). It is seen that only under the condition of constant driving force is the initial rise of the electrical PD, and hence of the total protonmotive force, high enough to make pumping power available almost instantaneously after the start of the pump. It also is seen that the pump is more stable under constant pumping rate, because after the pump is turned off, the PMF drops almost instantaneously in the lower curve whereas it declines gradually in the upper curve. Reproduced, with permission, from Ref. 11. Copyright 1982, Academic Press, Inc. H

H


14.

Ion

HEINZ

Pumps

329

a m e t h o d that a l l o w s r a p i d m o n i t o r i n g o f s u c h c h a n g e s b a s e d o n a c h a n g e i n l i g h t a b s o r p t i o n o f the n a t u r a l l y p r e s e n t s u b s t a n c e ,

bacteriorhodopsm,

h a s b e e n d e v i s e d f o r the l i g h t - d r i v e n p r o t o n p u m p s i n h a l o b i u m (12 ). B y this m e t h o d , i t c o u l d be shown with envelope v e s i c l e s of this b a c t e r i u m , that a f t e r i n i t i a t i n g the p u m p b y i l l u m i n a t i o n , the e l e c t r i c a l p o t e n t i a l (or the p r o t o n m o t i v e f o r c e ) r i s e s w i t h a h a l f t i m e of a b o u t 2 0 m s e c to a v a l u e c l o s e to 2 0 0 m v e v e n t h o u g h the b u f f e r c a p a c i t i e s o f a l l p e r m e a n t i o n s i n s i d e a n d o u t s i d e was k e p t h i g h e n o u g h to p r e c l u d e a p p r e c i a b l e c h a n g e s i n i o n d i s t r i b u t i o n d u r i n g that t i m e (13 ).

This

r e s u l t c l o s e l y a g r e e s w i t h what h a s b e e n p r e d i c t e d o n the b a s i s of the


c o n s t a n t d r i v i n g f o r c e m o d e l , a n d t h u s c o n f i r m s the h y p o t h e s i s o f the c r u c i a l r o l e o f the e l e c t r i c a l p o t e n t i a l f o r the e x p e d i t i o u s n e s s o f the coupling system. T h i s r e s u l t a l s o c o n f i r m s that the p u m p i s e l e c t r o g e n i c ,

accor

d i n g to the f o l l o w i n g e q u a t i o n d e r i v e d i n t e r m s o f n o n - e q u i l i b r i u m thermodynamics

(14):

a r e leakage coefficients of H

and Κ , respectively.

and

a r e s t o i c h i o m e t r i c c o e f f i c i e n t s of the p u m p f o r the i o n s i n d i c a t e d .

It i s

s e e n that a t c o n s t a n t c h e m i c a l p o t e n t i a l s o f the i o n s c o n c e r n e d the p a r t i a l d i f f e r e n t i a l i s d i f f e r e n t f r o m z e r o o n l y i n the p r e s e n c e o f a n e l e c t r o g e n i c p u m p , i . e. i f

Φ

Though buffer c a p a c i t i e s of

a l l p e r m e a n t i o n s c o n c e r n e d w e r e k e p t h i g h e n o u g h to p r e v e n t the f o r m a t i o n o f a n y s i g n i f i c a n t i o n g r a d i e n t d u r i n g the t i m e o f o b s e r v a t i o n a d i s t i n c t e l e c t r i c r e s p o n s e was r e g i s t e r e d u p o n i l l u m i n a t i o n a f t e r a few m i l l i s e c o n d s a s w o u l d b e i n c o n s i s t e n t w i t h a n e l e c t r i c a l l y s i l e n t H /K pump +

+

( i T

R

=

JT ). K

W h e t h e r the'higfr e x p e d i t i o u s n e s s r e s u l t i n g f r o m the e l e c t r o g e n i c P D r i s e w o u l d b e a n a d v a n t a g e f o r a n y s y s t e m i s h a r d to p r e d i c t .

Owing

to the l o w e l e c t r i c m e m b r a n e c a p a c i t y the i o n m o t i v e f o r c e at t h i s s t a g e , being predominantly electrogenic i s v e r y sensitive towards changing driving force.

the

B y contrast a m o r e slowly developing ionmotive force

g e n e r a t e d b y i o n t r a n s l o c a t i o n i s c e r t a i n l y m o r e s t a b l e a s the

energy

s t o r e d i n i t w o u l d s t i l l b e a v a i l a b l e f o r s o m e t i m e a f t e r the p u m p h a s stopped.

If the s y s t e m w e r e t o s u p p l y e n e r g y - r i c h c o m p o u n d s a t a

s t e a d y r a t e i n s p i t e of h i g h l y f l u c t u a t i n g e n e r g y i n p u t , o b v i o u s l y the s l o w e r but m o r e stable a r r a n g e m e n t would be m o r e suitable.

If,

o n the

o t h e r h a n d , the s y s t e m w e r e to a d j u s t i t s a c t i v i t y to a r a p i d l y c h a n g i n g r e q u i r e m e n t at a r a t h e r steady input of e n e r g y , but l e s s stable a r r a n g e m e n t would be m o r e

the m o r e

expeditious

suitable.


BIOCHEMICAL

330

ENGINEERING

O n the o t h e r h a n d , i t i s v e r y l i k e l y that the e l e c t r o g e n i c P D of a h i g h l y e x p e d i t i o u s c o u p l i n g s y s t e m a p p e a r s o n l y a s a wave of r a t h e r s h o r t d u r a t i o n , b e i n g r e p l a c e d l a t e r b y the c h e m i c a l P D ,

which m a y

p r e d o m i n a t e a f t e r the s y s t e m h a s r e a c h e d a s t a t i o n a r y state ( F i g .

2).

S u c h i s to be e x p e c t e d i f the " b u f f e r c a p a c i t i e s " of the p a s s i v e i o n s , c h i e f l y K - i o n s , a r e b i g e n o u g h to p r e v e n t the f o r m a t i o n of a m o r e stable diffusion potential.

H e n c e the p r o t o n m o t i v e f o r c e g e n e r a t e d b y

s u c h a s y s t e m m a y h a v e a d i f f e r e n t c o m p o s i t i o n i n the s t a t i o n a r y state f r o m that d u r i n g the i n i t i a l e l e c t r i c a l p o t e n t i a l r i s e .

In t h i s way,

the

a d v a n t a g e of a h i g h e x p e d i t i o u s n e s s at the b e g i n n i n g m a y be c o m b i n e d


with that of a h i g h e r s t a b i l i t y o b t a i n e d a f t e r s o m e t i m e of c o n t i n u o u s pumping activity.

F i g 2. a l s o s h o w s that the d e g r e e of s t a b i l i t y at a n y

s t a g e s h o u l d b e c o m e a p p a r e n t i f the p u m p i s s u d d e n l y s t o p p e d .

A f t e r the

p u m p h a s r e a c h e d i t s (stable) s t a t i o n a r y s t a t e , the p r o t o n m o t i v e f o r c e at that s t a g e would

decay only s l o w l y and

would t h e r e f o r e be

i m m e d i a t e l y a f t e r the p u m p h a s s t o p p e d .

measurable

In the i n i t i a l s t a g e ,

when the e l e c t r o g e n i c c o m p o n e n t of the P M F d o m i n a t e s ,

however,

the P M F

s h o u l d i m m e d i a t e l y d i s a p p e a r a f t e r s t o p p i n g the p u m p , a n d thus e s c a p e detection.

T h i s m a y b e the r e a s o n why the p r o t o n m o t i v e f o r c e h a s often

b e e n found i n a d e q u a t e t o s u p p o r t the c h e m i o s m o t i c h y p o t h e s i s o f o x i d a t i v e p h o s p h o r y l a t i o n , a s the e l e c t r o g e n i c c o m p o n e n t i s too l a b i l e to s u r v i v e the d i s t u r b a n c e a s s o c i a t e d with the u s u a l m e a s u r e m e n t s

of

electrical potential. Acknowledgment T h e s e studies were supported by a U S P H S - N I H grant No. R O I G M 26554-01. Literature Cited

1.

Heinz, Ε., "Mechanics and Energetics of Biological Transport"; Springer V e r l a g : Heidelberg, New

York,

2.

M i t c h e l l , P.,

144-8

Nature, 1961,

3.

Maloney, P.C.,

4.

Junge, W.,

5.

Witt, Η. Τ.,

6.

Stoeckenius, W.,

Wilson, T.H.,

Ber.

Mitchell, P.,

London,

1975,

Biochim. Biophys. Acta, 1979, Lozier, 505,

R.H.,

1978

J. Membrane B i o l , 1975,

Dtsch, Bot. Ges.,

Biophys. Acta, 1979, 7.

191,

25,

505,

355-427

Bogomolni, R.A.,

Biochim.

215

"Chemiosmotic Coupling i n Oxidative and

Photo-

synthetic Phosphorylation"; Glynn Research Ltd. : Bodmin, 8.

Mitchell, P.,

Skulachev, V. P., Ernster,

L.,

1966

"Chemiosmotic Coupling and Energy Transduction";

Glynn R e s e a r c h Ltd. : Bodmin, 9.

285

88, 283-301

1968

"Membrane Energetics"; Lee,

Eds; Addison-Wesley; p.

C. P., Schatz,

373


G.,

14.

HEINZ

Ion

Pumps

10.

Heinz, E., "Electrogenic and E l e c t r i c a l l y Silent Proton Pumps in Hydrogen Ion Transport i n Epithelia"; Shultz, I., Sachs, G., Forte, J.G., U l l r i c h , K.J., Eds; E l s e v i e r / N o r t h Holland Biomedical, 1980, p. 41

11.

Heinz, E., "Current Topics i n Membranes and Transport"; Kleinzeller, Α . , Bronner, F., Eds; Academic, 1982; V o l 16, p.249 Dancshazy, Ζ., Helgerson, S. L., Stoeckenius, W., i n preparation Helgerson, S. L., Dancshazy, Z., Stoeckenius, W., Heinz, E.; (Abstract); Biophys. J . , 1982, 37, 266a

12. 13. 14.

Heinz, Ε . , " E l e c t r i c a l Potentials i n Biological Membrane T r a n s port"; (Monograph); Springer, 1981


331

R E C E I V E D June 29,

1982


Ion Pumps as Energy Transducers - ACS Symposium Series (ACS

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