Cohn, H u
/
Isotopic lXOShifts in
3 1P
N M R of Adenine ivucleotides
913
Isotopic l 8 0 Shifts in 31PNMR of Adenine Nucleotides Synthesized with l80in Various Positions Mildred Cohn* and Angela Hu Contribution f r o m the Department of Biocheniisrry and Biophysics, Unicersity of Pennsylcania School of Medicine/G3, Philadelphia. Penns.vIcania 19104. Receiced Julj, 23, I979
Abstract: The analysih of phosphate IXOlabeled A D P and ATP by 3 ' P N M R spectroscopy at 145.7 and 235 M H z is described. The following isotopically labeled adenine nucleotide species were synthesized in various enzqme-catalyzed reactions: ADP!% PlXO3, A T P p - P 1 X 0 3 .ADPP-PlX04. A T P P - P ' x O ~ ( ~ - P ' X OAl T ) ,P P - P 1 X 0 3 ( y P ' X O ~and ) . ATP(P-P. y-P)lXOh.The magnitude of the isotopic (I8O)shift of the " P resonance varies from 0.01 66 to 0.0285 ppm in the p - P bridge and nonbridge positions and these resonances which differ by 0.01 18 ppm are completely resolved at 235 M H z . The y - P ( I X O )species are shifted b> 0.0220 ppm per IxOin both A T P y - P I 8 0 3 (three nonbridge) and A T P y - I X O(one ~ bridge. three nonbridge). For the latter compound. there was no indication of a difference in the chemical shift due to a bridge vs. a nonbridge I8O even at 235 MHz.
It has been observed'.* that I8O directly bonded to phosphorus causes a small shift of the order of 0.02 ppm in the chemical shift of "P. At high frequency, e.g., 145.7 MHz, the '80/160 in inorganic phosphate can be determined with considerable accuracy a t high values of lXO.'Even at lower frequency, 40 MHz, in spite of the lack of resolution, good 0 between 3 ' P N M R and mass agreement of ' 8 0 / ' hratios spectrometric analysis was achieved by using a curve a n a l y ~ e r . ~ I n the current study, the lXOanalysis by 3 1 PN M R has been extended to A D P and ATP isotopically labeled in various positions of the molecule and the correlation of the magnitude of the isotopic shift to the double-bond character of the P - 0 bond was examined.
Experimental Section K H ~ P O was J exchanged thice with H2IXO(95.8% IXOenrichment) to yield a product containing 93.6% 180.4 Carbamate kinase, PEP.' A W P , and A T P were purchased from Sigma Chemical Co. and A D P u a s purchased from Boehringer Mannheim. K C N O was obtained from Fisher Scientific Co. MgClz (99.999% pure) was a product of ;\ccurate Chemical and Scientific Corp. IXO-Enriched Mater w a s purchased from Miles. Pqruvate kinase was a gift from Dr. George Reed of the Cniversitl of Penns>lvania and adenqlate kinase u a b a gift from Dr. Noda Lafabette of Dartmouth Medical School. TEA from Eastman M ~ redistilled S before use. PEI plates Mere purchaaed from Brinkmann. All other reagents used were reagent grade without further purification. The composition of all s)nthesized I8O nucleotides was checked on a PEI plate developed lrith I .5 .M LiCI. N M R Measurements. The 3 1 PN M R spectra were recorded at 145.7 M H 7 on a Bruker W H 3 6 0 N M R spectrometer: samples of 1.5 m L i n IO-" N M R tubes maintained a t 25 O C mere used. One sample \ \ a s measured at 235 MH7 on the Carnegie-Mellon 600-MH7 spectrometer i n a 5-mm sample tube a t I 9 "C. Synthesis of ATP-y1*03(11). The A T P ( y P - ' X 0 3 )( 1 1 ) w'is thcsized in two steps bq a modification of the procedure used t o . thesize Y - ~ ~ P - A T PI-Carbamyl ." phosphate is synthesized chemically from P,(lXO)and K C N O and the l X Ophoaphorbl group in the carbamyl phosphate produced is transferred to A D P to form A T P ( y P AMP + A T P ( t r a c e a m o u n t )
lXO3)(11) \+ith three nonbridge lXOatoms in a reaction catalyzed by ca r ba nia t e k i n ase .' KH2PIXO4(300 p L of 1 M ) (93.6% lxO). 300 p L of 1 M sodium acetate. pH 4.8, and 375 p L of 6 hl K C \ O (prepared immediat before use) uere incubated in a hater bath a t 30 O C . The pH \ monitored at 5 - m i n intervals: acetic acid \+as added to maintain the p t l at about 6.5. A n additional 100 p L of KCKO was added to the mixture after 30 min and incubated at pH 6.5 for a n additional 15 min. The reaction mixture was then chilled in ice. T o the cold mixture Mere added 550 p L of 550 m M ADP. pH 7.6, I .5 mL of 1 M Tris-HCI. pH . 30 units of carbamate kinase in 50 7.3. 300 p L of I bl M ~ S O Jand p L . The tnixture ua5 incubated a t 38 O C for 30 n i i n . The yield of A T P - y l 8 0 3 from A D P u a s 95%: the unreacted 5% was mainly due to the contamination of A M P in the A D P used. On the basis of [ '80]orthophosphate. t h e yield was 76%. The reaction was stopped by the addition of 450 p L of 50% trichloroacetic acid: the mixture \vas centrifuged. and the supernatant ~ L I extracted S thrce times with five volumes of ether. The components of the reaction mixture were separated on il dieth~laminoethqlccllulose(DE-52) column ( 3 X 25 cm). The adenine nucleotides were eluted \rith a 4-L gradieni, 0-0.5 M , of TEA-CO: buffer. pH 7.6 (buffer w a s prepared by bubbling CO: into a T E A solution). PI \\as eluted bet\\een 0.165 and 0.205 M salt, A M P at 0.265 M. A D P at 0.285 M, and A T P a t 0.35 M salt. All three peclks \%ere\+ell resolved from one another. After the ATP fractions here concentrated i n a rotary evaporator to near drkness. the A T P \+:IS diswlvcd in a feu milliliters of H 2 0 and titrated to pH 8-9. The metal ion\ \\ere removed from A T P bq extraction v,ith 1 % X-hydroxyquinoline.' The P,-I&Ocould also be recouercd. Sqnthesis of ADP-BIs03 ( I ) and .ATP-fi1*03,y1*0 (VI).The isotopically labeled A T P and A D P were bynthesized from isotopically labeled carbamyl phosphate, A M P , a trace of ATP, and the en7ymes adenqlate kinase and carbamate kinase as indicated schematically i n eq 1-4. KH2P"Od (20 mg). I mL of I L1 sodium acetate. pH 4.8. and I ,25 iiiL of 6 M K C N O were incubated as described above. After 25 min. 0.4 niL of 6 M K C l O \ a s added and incubation u a s continued for I5 min. The mixture was then chilled in ice. To the cold misture \\ere added the following: 500 pmol of A M P in 0.875 mL. 8 pmol of A T P in 0.6 mL, 0.6 mL of I M MgCI;. 5 mL of I M Tris-HCI. pH 7.3. and
-
adenylate kinase
-
2ADP
(1)
carbamate kinase
* A T P ( Y P ' ~ O +~ )CO:
ADP + carbamyl-P("O,) adenylate kinase
AMP + A T P ( y P I B O , )
.
A D P ( O P 1 8 0 0 ,+) c a r b a m y l - P ( l8O0,)
-
carbamate kinase
000'-7863,%0/
1502-0913S01 00/0
(2)
ADP + ADP(OP"0,) three nonbridge 0 atoms 0 0 0
II
* A T P ( P - ? . - P 1 8 0 , ) A--O-P-O-P-+P-O
adenylate kinase
+ NH,
l 0
I1 0
c 1980 American Chemical S o c i c t ~
(3)
II
I
0
(4
914
Journal of the American Chemical Society
/ I02:3 /
January 30, 1980
Table 1. 3 ' P Chemical Shift (ppm) (Upfield) per I8O Bonded to P Atoms of A D P and A T P (B = Bridge. N B = Nonbridge) nucleotide species
P
CY
7
e I.
II
ADP.8 '0 AMP-0-P-0
0.0220
I 0 0 I1
ATP-71'0
0
II
I/
AMP-0-P-0-P-0
I 0
0.0220
l 0
0
LII.
I/
ADP B QOIAMP-0-P-0
0
0 IV
ATP
II
I/
AMP-O-P-+P-O
I 0
I 0 0
V
ATPp"0, yL'O AMP-0-P-0-P-0
II
I
0
0
0
0
II
I/
0.0220
0.028 1 ( S B) 0.0165 (P-7 B)
0.0220
0.0285 (NB)O 0.0163 (B)"
ATP@ -0,710, AMP-0-P-O-P-•
I
VII.
0.0165 (P-7 B) 0
II
I
VI
0.02 1 5
0.0166 (CY-PB)
I 0
/
0 0
0
II
II I
(0-7 B)
0.02 I 7
0
.4TP@dl'0,,n"01 .4MP-O-P-O-P-O
I
0.01 72 (CY-@ B)
0.0281 (NB) 0.0165 (CY-pB)
~
(' Determined from resolved peaks a t 235 M H z , all other bridge and nonbrtdge values were estimated from spectra dt 145 7 M H 7 15 m M ADP-PlX03,and 25 m M P-enolpyruvate, 0.4 mgof pyruvate kinase was added. The reaction mixture was incubated at 25 " C for 30 min. The reaction was stopped by diluting the mixture with 0.2 M T E A - C O 2 buffer, pH 7.6. The mixture was applied to a 3 X 25 cm DE-52 column and was eluted with a 2.2-L gradient, 0.2-0.45 M of T E A - C O l buffer. pH 7.6. A T P - P ' Y 0 3 y ' 8 0 1was eluted at 0.35 M
0.6-
-
0 W
N 4
om4-
0
002
0 I I
tI
I
I
A--O-P-O--P-O
-
0 0#07 0
T
I
IO
I
1
20
I
I
30
T i m e (min) Figure 1. Distribution of nucleotides as a function of time in the preparation of A D P P ' X 0 3( I ) and ATPP-. y - l x 0 h ( V I ) .
2.5 mL of 1 M KCI. The reactions were initiated with 1500 units of adenylate kinase (0.I5 mL) and 50 units of carbamate kinase (0.1 mL) and the reaction mixture was incubated at 38 'C for 30 min. The course of the nucleotide concentrations over the 30-min period is hhown in Figure I , The mixture uas chilled in ice immediately and cold HCIOJ w'as added to a final concentration of 6%. After removal of the precipitated protein by centrifugation, the supernatant was titrated to pH 7.6 w i t h K O H . After KClOl was allowed to precipitate at 0 O C for at least 30 min. the precipitate was centrifuged and the supcrnatant was applied to a DE-52 column. The column was eluted k\ith 3 1. of TEA-CO2 buffer, pH 7.6, gradient 0-0.5 M. The fractions containing ADP-@lX03( I ) and ATP(P,y)-IX06( V I ) , wpcctively, were collected, and. after the solutions were concentrated w i t h ii rotary evaporator to near dryness, metals were removed from / \ T P ( V I ) a s described above. A n A D P - P I X 0 3solution at pH 8.0 was applied to a 1.9 X 5 cm column of acid-washed Chelex IO0 to remove l l l C tal s. Synthesis of ATP-BPIR03( V )(B-7 Bridge I8O).To 6 mL of solution containing 100 niM Hepes, pH 7.6, 25 m M MgSOJ, 100 m M KCI,
+ PEP
pyruvate kinase
pyruvate
i 0
I + A--O-P-O-P-O--P--O I 0
0
I
I
0
0
I I 0
salt. The sample was purified before storage as described above. Conversion of ATP-ylX03(11) to ATP-B1*03(VII) (One CY-@ Bridge, Two Nonbridgel. A solution containing 20 m M Hepes buffer, pH 8.5. treated with Chelex, 8.3 m M A T P - 7 I 8 0 3 ( I I ) , 2 m M valine treated with Chelex, 15 m M MgC12, and 1 m M ethylenediaminetetraacetate in I .5 m L of 20% DzO was incubated at 25 'C. The partial reaction shown (eq 5) was initiated by the addition of I8 ~g of valine t R N A synthetase and the time coursc was monitored by the appearance of l X Oin the C Y - ~ peak. 'P At equilibrium there are three y - P species I , l X O ~ ' hand O ~0.508 , two a - P species (0.354 1 x 0 3 1 x O0.138 (0.445 1 8 0 1 ' 6 0 3 and 0.555 and, four p - P species (0.354 " o b i 8 0 2 n b i 6 0 1 , 0.092 ' 8 0 b ' 8 0 n b 1 6 0 ~0.046 , i 8 0 2 n b i 6 0 2 , p and 0.508 " 0 4 where the subscript b indicates bridge and nb, nonbridge). The initial spectrum and the spectrum at equilibrium are shown in Figure 2.
Results Isotopic ('*O)Shifts of 31PResonance. B a s e d on t h e statistical distribution, for ATP-? I8O3 ( I 1) containing 89% I8O,four species exist (0.704 ' 8 0 3 1 h 0 1 , 0.26 1XOz'h02, 0.032 ' X O ' h 0 3 , and 0.001 3 I6Oj);however, only t h e first t w o species could b e
1 Isotopic
Cohn, H u
3 iP
I8O S h i f t s in
N M R of Adenine Nucleotides
EOU~L~BR~UM
I/
1 1
1
915
(IbJnb)
\
I
qz
, .'
ia
1 Fin.' uL-4
I
_-
-*-
Figure 2. 3 i PN M R spectra of the conversion of ATP?l8O3( I I ) , 89% I8O, to ATP8-'803( V I I ) The upper spectrum is the initial spectrum of a 1 5-mL solution containing 20% DzO, 8 3 mM ATP, 2 m M valine i n 20 m M Hepes buffer pH 8 8, 1 m M EDTA The lower rpectrum was recorded 7 h after the addition of 13 m M MgClz and 18 ~g of Val41 t R \ A sJnthetase of Escherich/a colr, 40 m'M EDTA was added before the spectrum was recorded N M R parameters 145 7 MHz, 100 scan?. acquisition time 4 5 s, 45' f l ~ pangle ap =
( a , b)
0
0
a =Numberbf innontvidgeposition b :Number of "0 's in bridge position
0
1
0
4\\ v a l y l AMP
'I
+E
i 0
I
A--O-P-O-P-0-P-0
I
0
0
I I 0
I I 0
O--P-*P-O
0
I
valyl AMP
+E
I -.P-*P-O I 0
1
1
0
I I 0
0
I
1
1
- 5500
0
0
I I 0
A-0-P-0-P-0-P-0
1
1
1
Hz from PO,
(5)
determined quantitatively with the sensitivity available as shown in the upper spectrum of Figure 2. The chemical shift per I8O is 0.0220 ppm (cf. Table I ) . Analogous spectra were obtained for all isotopically labeled nucleotides listed in Table I , two A D P species ( I , p - i 8 0 3and III,p-~x04), and five A T P species (Il,y-''O3, I V , y - i 8 0 j , V,P-IXO3(p-y bridge), VI, ;%y-i806,and VII,p-i803(a-i3bridge)). The syntheses of these compounds are described in the Experimental Section with the exceptions of A D P I l l , which has been previously described,' and A T P IV, which was synthesized chemicallq by Dr. M. R . Webb. The possibility of resolving peaks for the p-P with equal numbers of atoms differing only in the bridge vs. the nonbridge position first appeared feasible from the lower spectrum shown in Figure 2, an equilibrium mixture of A T P I I and A T P VI1 where two overlapping 3 1 Presonances for the p - i 8 0 2species were observed from A T P VII. T o obtain a resolved spectrum, where the species with lYOlas well as IxO* could be observed and resolved for both p - P and y - P of ATP, A T P VI, /3-, y-I8O6 was synthesized with -63% I8O as described in the Experimental Section and recorded at 235 MHz. The spectrum of the 13-P of A T P VI is shown in Figure 3 and two types of I8O1,and of I8O2and lXO3 all six lines due to are resolved; the chemical shifts are listed in Table I . Discussion The observable chemical shift of the ) ' P resonances due to lXOsubstitution in the phosphate group is a powerful tool to solve many mechanistic aspects of phosphate reactions.i,3,xI I However, the shift can be determined directly a t high frequency only, e.g., a t 145.7 M H z or even more readily a t 235
I
1
-5525
I
I
I
I
I
-
-5550
-3
Figure3. " P I \ l ~ R s p e c t r u n i o f t h e B - P o f - 6 3 % 1XOATP-P-.y1806(VI) recorded t t 23.5 MH7 by correlation \pectroscopj 29 000 scans. sweep time I \
MHz. Lower frequency instruments (40.5 MHz) may be used in conjunction with a curve a n a l y ~ e rSome . ~ problems emerge at high frequency which do not occur at low frequency. In particular, chemical exchange effects among various species cause greater line broadening. For example, the exchange rate between various metal chelates of A T P or with free A T P will affect the line width not only of the y-P, but also of the CY- and p-P. With the most commonly used metal ion, Mg*++,the line width of the 3 i Presonance at 24.3 M H z is 0.9 H z for both MgATP and free ATP. At 145.7 MHz, the line width of metal free ATP is 1.2 Hz, but, after addition of a saturating amount of MgC12 (99.999% pure), the line width of the a - P increased to 2 Hz and the line width of p- and y - P increased to 4 H z undoubtedly due to exchange among the various MgATP species in solution. I f Mg'+ is substituted by Ca'+, with exchange rates -1000 times faster than those of Mg2+, no such line broadening is observed. For the most part, the magnitude of the IxOshift on the 3 1 P resonance correlates well with the amount of double-bond character of the bond as indicated in Table 11, the difference being greatest between the shifts of p - P of A T P due to the nonbridge oxygens (0.0285 ppm) and the c - p or 8-7 bridge oxygen (-0.01 67 ppni). A similar difference between the effect due to bridge and nonbridge oxygen has been reported for [ a p - i X O ] A T P a Sand [ c v - ' ~ O ] A T P ~ SHowever, .'' the shift on p - P due to the two nonbridge oxygens of the 6-phosphate of A T P shows a considerable deviation from the calculated value. The differences due to the bridge oxygen between the pairs A D P f l ' x 0 3 - A D P p i X O jand A T P y l X 0 3 - A T P i X 0 are 4 apparently too small to detect at 145.7 MHz. It is somewhat
Journal of the Anzrrican C'hrmical Societj,
916 Table I I . ' I P C h e m i c i l S h i f t per l X O'15 Tq pe, ,2 '' source
of " 0
,V
phosphoanhydride bridge ( A D P , A T P ) phosphate anion t c r i n i n a l - P i x 0 3 anion ( A D P , A T P ) A T P 6-nonbridee l X O
+
Function o f P 0 Bond
1.00 1.25 1.33 1.50
shift, p p m calcdh obbd (av)
0.0208
0.0 166' 0.0206
0.0221
0.0220
0.0249
0.0285
+
102:3
/
Januarjs 30, 1980
Acknowledgments. The authors wish to thank Dr. M. R. Webb of the University of Pennsylvania for making his " P N M R data on ATP i x 0 4 available before publication and Drs. A. A . Bothner-By and J . Dadok of the Carnegie-Mellon N M R facility for obtaining the ? ' P N M R spectrum at 235 MHz. This work was supported in part by L S P H S Grant G M 12446 and NSFGrant PCM78-13633. N M R spectra at 145.7 M H z were recorded at the Middle Atlantic N M R facility supported by U S P H S Grant RR542 at the University of Pennsylvania.
N = E(la 2 b ) / ( a b ) where a = number of single bonds and h = number of double bonds. Calculated f r o m the value observed for .V = I . Averageof31Pshiftsduetothea-flbrldge'800nru-3iP References and Notes a n d p - 3 1 Pa n d to the 6-7bridge lXOon p - 3 1 P .
surprising, however, that even a t 235 MHz where the P-r bridge l8Oand nonbridge I8O species on p - P of ATPP,y-180h are well resolved (see Figure 3), the spectrum of the y-P(I8O4) gave no indication of any nonequivalence of the shifts due to the nonbridge l X Oas compared to the P-y bridge lXO.A l80?, simple five-line pattern corresponding to l 8 0 o , 1801, lXO3,and " 0 4 was observed for each of the two regions of the y - P doublet of the 63% I8Osample at 235 MHz. T h e equivalence within experimental error of the chemical shift of the bridge and nonbridge oxygens on the y-phosphate of ATP and the nonequivalence for the P-phosphate of ATP may be related to the axial symmetry in the chemical-shift tensors of the dianionic forms of mononucleotides and the axial asqmmetrq for the anionic form of diesters.I4 Since the chemical-shift tensor of the acid form of mononucleotides is no longer axially symmetric, it would be of interest to examine ATP lXO4in the acid form.
Cohn. M.; Hu, A. Proc. Nafl. Acad. Sci. U.S.A. 1978, 75, 200-203. Lutz. 0.; Nolle, A . ; Staschewski. D. Z. Naturforsch. A 1978, 33, 380382. Van Etten, R. L.; Risley, J. Proc. Nafl. Acad. Sci. U.S.A. 1978, 47844787. Cohn, M.; Drysdale, G. J. Biol. Chsm. 1955, 276, 831-846. The following abbreviations are used: PEP, phosphoenolpyruvate; TEA, triethylamine; PEI, polyethylenimine; Hepes. 4-[2-hydroxyethyl)-l-piperazineethanesulfonic acid. Mokrosch, L. C . ; Caravaca, J.; Grisolia, S. Biochim. Biophys. Acta 1960, 37, 442-447. Metzenberg, R. L.; Marshall, M.; Cohen. P. P.;Miller, W. G. J. BiOl. Chem. 1959, 234, 1534-1537. Brown, F. F.; Campbell, I. D.; Henson. R.: Hurst, C. W. J.; Richards. R . E. Eur. J. Biochem. 1973, 38, 54-58. Webb, M. R.; McDonald, G. G.; Trentham, D. R. J. Biol. Chem. 1978, 253, 2908-291 1. Bock, J. L.: Cohn, M. J. Bioi. Chem. 1978, 253, 4082-4085. Lowe, G . ; Sproat, B. S . J. Chem. Soc., Perkin Trans. 7 1979, 16221630. Jordan. F.; Patrick, J. A.; Salamone, S., Jr. J. Biol. Chem. 1979, 254, 2384-2386 Jarvest R L Lowe G J Chem S O C , Chem Commun 1979, 364366 Terao, T Matsui S Akasi, K J Am Chem SOC 1977, 99, 61366138
Nucleoside Complexing. A Raman and 13CN M R Spectroscopic Study of the Binding of Hard and Soft Metal Species Luigi G . Marzilli,* Baltazar de Castro, John P. Caradonna, Robert Charles Stewart, and C. P. Van Vuuren Contribution f r o m the Department of'C'heniistrj.. The Johns Hopkins L:nirersitj, Baltimore, Maryland 21218. Receirrd March 16, 1979
Abstract: T h e influence o f a u i d e variety of metal salts a n d complexcs on the R a n l a n and I ' C \ M R spectra of three o f thc four c o m m o n nucleosides (uridine, adenosine, and cytidine) in M e r S O has been determined. ScLcral related molecules also htudied included deoxycytidine, 5-methyldeoxycytidine, 8-bromoadenosine. 6-dimethylaminopurine-9-riboside. a n d .VJ..VJ-dimethq Ia m i n o - ] -methylcytosine. Inorganic species utilized included cis- [ P t ( Me2SO):C12]. HgCI:.Zn(\03)2, C d ( Y O 3 ) 2 , Pb(N03):. Pb(C104h PbC12, Mg(N03)2, C a ( N 0 3 ) ~S r ( N 0 4 2 . B a ( N 0 3 ) 2 . BaCI:. La(KO3)l. Pr(\O3)3. L U ( N 0 3 ) 3 . LU(CIOJ)?, G L I ( N O ? ) LiCIO4, ~, N E t J C I , and N E t 4 h 0 3 . 'The I3C h M R shift dependcncieh on nictiil concentration were used to c;iIcuI;itc f o r m a t i o n constants for selected purine and p y r i m i d i n e derivatives. Constants obtaincd \ + i t h tigC12 agreed w e l l u i t h literature v;iIues. Hovdever, o u r studies suggest t h a t alkaline earth m e t a l ions have ii lower a f f i n i t y for nucleosides t h a n t h a t suggested previouslq. Of the c o m m o n nucleosides. cytidine has the greatest affinit! for ;ilkalinc c,irth mctal ions and these nppcar to b i n d to 0 - 2 . R a n i a n and I3C N i M R studies w i t h N 4 , N 4 - d i m e t h y l a m i n o - I-methqlcytosinc arc presented \\ h i c h support the 0 - 2 b i n d i n g mode. T h e only electrophile capable of interacting w i t h N-3 of this p q r i m i d i n c i h the H + ion. Conclusions d r a u n about b i n d i n g sites to other nucleosides are i n agreement w i t h previous studie5. A \ u m m a r q kiblc specifking the importancc of anion and cation b i n d i n g to the four common nucleosides is included. This s u i i i m a r ) providch ;I riitioniilc for ;ill oi'thc literature obscrvations a n d should help to m i n i m i z e the confusion currently surrounding the n'iturc oi' the iiitcractlons ot'inorganic hpccics u i t h nucleosides in Me?SO.
Introduction Growing recognition of the importance of metal ion interactions with nucleic acids and nucleotides has stimulated an 000~-7863/80/l502-0916901. 0 0 / 0
explosive growth i n the number of studies devoted to understanding the chemistry of the complexes formed. This interest has Icd to at least seven recent reviews of this topic.' The great promise of platinum( I I ) metalloantineoplastic
'
'c' 19x0 A t n c r i c a n C h c m i c a l Socict)
