J. Phys. Chem. 1992,96, 10036-10043
10036
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Pulse Radiolysis Studles of Intramolecular Electron Transfer In Model Peptides and Tyro Radical Transformation In H-Trp(Pro),-Tyr-OH Series of Proteins. 5. Trp* Peptldes Krzysztof Bobrowski,+ Jerzy Holcmaqt Jaroslaw Poznaiiski,+Marek Ciurak,s and Kazimierz L. Wierzcbowski**+ Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Rakowiecka 36, 02-532 Warszawa, Poland; Department of Environmental Sciences, Riso National Laboratory, DK-4OOO Roskilde. Denmark; and Institute of Chemistry, University of Gdahk, Sobieskiego 18, 80-592 GdanSk, Poland (Received: May 29, 1992; In Final Form: August 13, 1992)
The kinetics of intramolecular long-range electron transfer (LRET) between neutral tryptophan radical and tyrosine in aqueous solution of H-Trp(Pro),-Tyr-OH, n = 4 and 5 , peptides has been studied by pulse radiolysis over the temperature range 288-328 K. The rate constants, kl, and thermodynamic activation parameters of LRET thus obtained, together with those for shorter (n = 0-3)peptides of the same series, measured earlier, are analyzed in terms of Marcus nonadiabatic theory of LRET and distributionsof donoracceptor distances and angular orientationsdetermined by the conformational energy calculations. To explain the observed exponential decrease of k, with the number of n of Pro residues, simulation of the overall distance dependence of the rate according to models assuming involvement of LRET (i) through-bond (TB), (ii) through-space (TS), and also (iii) through both pathways simultaneously were performed by fitting the calculated mean rate constants, (kTB)and (kTs),to the experimental k , data. The best agreement between the experimental and calculated rates was obtained for a modified version of the last model (iii), according to which competitive electron transfer through the TS pathway occurs only in the conformers exhibiting van der Waals contacts and favorable angular orientation for a large overlap of T and u orbitals between the indole and phenol rings. The best-fit rate constants obtained indicate that in short-bridgedpeptides (n = 0-2)electron transfer takes predominantly the TS pathway, while in longer ones (n = 3-5) it occurs mainly by the TB pathway (BTB = 0.28 k 0.05 A-' at 298 K). Descriptors of electronic (@ = 0.2A-') and nuclear (AB= 0.1 A-I) contributions to the overall distance dependence of the TB-LRET were roughly estimated from the distance corrected for an expected thermal longitudinal expansion dependenceof activation parameters, derived from rate constants, kl,cor, of the oligoproline bridge.
Introduction The intramolecular oneelectron redox reaction (eq 1) in model peptides H-Try-(X),,-Tyr-OH, where X is an amino acid residue, involving tyrosine to tryptophan radical (Trp') electron transfer, H-Trp'-(X),,-Tyr[OH]-OH
kl
H-Trp[H]-(X),-Tyr'[O]-OH (1)
has been intensly studied by pulse radiolysis' and laser flash photolysis2 in several laboratories. The electron transfer in H20 solution close to neutrality is also accompanied by a net proton transfer due to initially formed TrpH'+ and TyrOH'+ (pK, of 4.3 and 0. We found that these conditions are fulfilled only for a narrow range of the thermal expansion coefficient, the values of which proved roughly proportional to the extent of helical ordering and thermal stability of the -(Pro),,-bridge>b+d a( Ar(CB))/6T:4 . 0 1 1 for 3 and 5, and derived -0.016 A n-’ K-’for 4. The activation enthalpy, AHHmr*, from temperature dependence of kl,,r varied linearly with n = 3-5 (Table 11). Assuming (Ar(CB))= 2.7 A n-I at 298 K and PTB= 0.28 A-I, we obtained 0.1 and /3JB= 0.2 A-I, respectively. Larger values of the thermal expansion coefficients led to the reversed order of AS*and negative sign of the coefficient
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Bobrowski et al. by DeFellipis et al.Ih and discussed also elsewhere,” on the basis of different values of the descriptor 3r, estimated for reaction 1 across H-Trp(Pro),-Tyr-OH and H-Tyr-(Pro),,-TrpOH, 0.37 and 0.23 A-I, respectively. In light of our findine, this difference is most likely due to somewhat different equilibrium populations of cis and trans isomers about X-Trp and X-Tyr bonds and XI rotamers of C- and N-terminal Trp indole side chain, resulting in a different average separation distance between the aromatic rings in the two peptide series. In a similar more rigid conformationally metallo system [(NH3),0s-isonicotinyl-(Pro),,-Ru(NH3)5]4+,24b numerical values of /3 descriptors were obtained using r values calculated from X-ray diffraction data for fibrous PLP I and I1 and 13CNMR data for cis-trans equilibrium in model series of compounds, v k , [(NH3),Co(Pro),,13. However, the latter are lacking N-substituted bulky isonicotinyl group, which in the electron-transfer Os-Iso-(Pro),,-Ru system studied may shift this equilibrium about Iso-Pro peptide bond toward the cis isomer. Therefore, the r and 8 values obtained24bmay prove somewhat different accordingly.
Conclusions We have shown that the experi,mentallyobserved exponential dependence of the rate of intramolecular LRET between tyrosine and tryptophan radical in peptides H-Trp(Pro)-Tyr-OH, n = 0-5, aTB on the number n of separating proline residues can be satisfactorily Pel ’ To verify the explanations presented, the temperature and time explained within the framework of Marcus theory, provided that dependent distribution of distances between various atoms of the conformational properties of the peptides studied are properly bridge and of aromatic rings should be determined from molecular accounted for. It was namely demonstrated that for determination dynamics calculations. As a next step, quantum mechanical of a physically meaningful value of the descriptor 8 of the overall calculations of the electron-tunneling matrix elements for the TB distance dependence of LRET, it is not sufficient to merely correlate according to eq 2 the experimental rates with the caland TS pathways might be also attempted. Until it is done, we culated mean increment per Pro residue of the separation distance are tempted to believe that the temperature-induced elongation between the redox pair along the peptide backbone, as is done of the -(Pro),,- bridge in longer peptides and temperature-deusually. Because short-bridged pepti& 0-2 were found to contain pendent hydrophobic clustering of indole and phenol rings can be considered the main cause of decrease of the experimental AH‘ a high population of conformers with close van der Waals contacts and A with n.33 between the side-chain indole and phenol rings, involvement of Comparison with Related Studla on -(Pro),,-Bridged LRET both the TB and TS pathways had to be included and respective Systems. The weak distance dependence of the nuclear factor relative rates of LRET for all conformers calculated. Averaging found in the system studied, compared with that reported for the of the latter over all low-energy conformers allowed to account properly for the contribution to the rate of LRET of individual analogously bridged system bearing charged metal ions:24 [(NH3)sOs-iso-(Pro),,-Ru(NH3)5]4+, seems to be consistent with species, proportional to their Boltman probability, the donorthe neutral character of the Trp’ and Tyr’ radicals involved in acceptor distance and, in the case of the TS-LRET, also to the LRET. The reorganization parameter A, calculated from AHm* donor-acceptor angular orientation. Fitting the calculated mean data and W = 4 . 1 1 eV by cq 6, varies only slightly with n from rates for the TS and TB pathways to the experimental rates -1.1 to -1.2 eV. In the system Os(II)/Ru(III), where nuclear showed that electron transfer along the TS pathway dominates contribution to the distance dependence of LRET has been shown in short peptides (0-2) and occurs through direct contacts between to predominate (8, = 0.95 and 8 = 1.6 A-I), the reorganization the aromatic rings; in longer peptides (3-5)it takes mainly the energy, A, rose with r from =1.0 eV (n = 1) to -1.5 eV (n = TB pathway. The experimental thermodynamic activation parameters of 3), Le., substantially more than predicted by an ellipsoidal cavity reaction 1 are probably implicit functions of activation of the mode1.z4a*34 To explain much weaker distance dependence of the rate of underlying TB and TS mechanisms of LRET and of thermally induced elongation of the -(Pro),,- bridge. Because of this it was LRET in our system than that observed in some other prolinepossible to estimate roughly only the electronic and nuclear bridged LRET ~ y s t e m s , 2we ~ , estimated ~~ the electronic coupling contributions to the overall distance dependence of the rate of matrix elements, HDA, between Trp’ and Tyr aromatic rings (Table corrected 11). The values of HDA,m, calculated by eqs 3-5 with use of X, TB-LRET, using AH*data for longer peptides (H), for an assumed longitudinal thermal expansion of the -(Pro),,and k,,, (298 K),are rather low and decrease with n (3 5) from 0.22 to 0.13 cm-I. The ratio HDA,,,/HDA,i = ( k l , , , / k l , l ) ~ ~ z ,bridge. Values of the corresponding descriptors (& > &) indicate ~alculated’~ from the experimental kl (298 K) data, varies with that the TB mechanism is largely under control of the electronic n (1-.c 5) from 1 to 0.1 1, in reasonable agreement with the order factor. The high value of the electronic coupling decay factor per of the absolute HD- values. The low values of H D A explain why the rates of LRET in the system studied are by about 2 orders peptide bond along a PLP I1 type helical backbone, cB = 0.86, found in this work, significantly higher than the average value of magnitude lower than those found across the same spacer for metal-teligand the Os(II)/Ru(III) r e d 0 x ~ or ~ 9dr(Re)/r*(bpy) ~ of this factor estimated” for proteins (ee = 0.6), suggests that charge-tran~fer~’ pairs, characterized by approximately 1 order helical segments in protein LRET systems can function as very of magnitude higher H D A values at the same n (measured for n efficient channels for electron transfer. 1-3). Despite the relatively low level of sophistification, the applied It is worth noting that the rate of LRET of the same order of models teach as very important lesson that estimation from exmagnitude as that observed for reaction 1, and similarly dependent perimental data for semirigid bridged systems of the overall distance dependenceof the rate of LRET and contributions thereto on the separation distance (Ar(C,)) with /3 = 0.37 A-I, has been foundB for oneelectron radical transformation Met/S.-.Br Tyr’ from the TB and TS pathways, on the one hand, and electronic in analogously bridged system H-Tyr-(Pro),,-Met-OH. and nudear factors, on the other, should be made with full account The apparent directional specificity of LRET was suggested of conformational properties of molecules studied.
@TB
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ET in Model Peptides Acknowledgment. This work was in part supported within the Projects CPBR 3.13 and CPBP 01-19. Financial support to K.B. from the Riso National Laboratory (Denmark) is appreciated.
References and Notes (1) (a) Priitz, W. A.; Land, E. J. In?. J . Radia?. Biol. 1979,36, 513. (b) PrUtz, W. A.; Land, E. J.; Sloper, R. W. J. Chem. Soc., Faraday Trans. I 1981, 77, 281. (c) Butler, J.; Land, E. J.; PrUtz, W. A.; Swallow, A. J. J . Chem. Soc., Chem. Commun. 1986, 348. (d) Butler, J.; Land, E. J.; Priitz, W. A.; Swallow, A. J. Biochim. Biophys. Acra 1982,705, 150. (e) Bobrowski, K.; Wimhowski, K. L.; Holcman, J.; Ciarak, M. Stud. Biophys. 1987,122, 23. (f) Faraggi, M.; DeFelippis, M.R.; Klapper, M. H. J . Am. Chem. SOC. 1989, 111, 5141. (8) Bobrowski, K.; Wierzchowski, K. L.; Holcman, J.; Ciurak, M.In?. J . Radia?.Mol. 1990,57,919. (h) DeFelippis, M. R.; Faraggi, M.;Klapper, M.H. J. Am. Chem. Soc. 1990,112, 5640. (i) Faraggi, M.; Klapper, M. H.In Excess Electrons in Dielectric Media; Ferradini, C., Jay-Perin, J.-P., Eds.;CRC Press: Boca Raton, FL, 1991; Chapter 13. (2) Sloper, R. W.; Land, E. J. Photochem. Phorobiol. 1980, 32, 687. (3) (a) Posner, M. A.; Adams, G. E.; Wardman, P.; Cundall, R. B. J . Chem. Soc., Faraday Trans. I 1976,72,2231. (b) Jovanovic, S.V.; Simic, M.G. J . Free Radical Biol. Med. 1985, I , 125. (c) Bent, D. V.; Hayon, E. J . Am. Chem. Soc. 1975,97,2612. (d) Dixon, W. T.; Murphy, D. J . Chem. Soc., Faraday Trans. 2 1976, 72, 1221. (4) (a) Priitz, W. A,; Butler, J.; Land, E. J.; Swallow, A. J. Biochem. Biophys. Res. Commun. 1980,96,408. (b) PrUtz, W. A.; Siebert, F.; Butler, J.; Land, E. J.; Menez, A.; Montenay-Garestier, T. Biochim. Biophys. Acra 1982, 705, 139. (c) Ghiron, C. A.; Santus, R.; Bazin, R.; Butler, J.; Land, E. J.; Swallow, A. J. Biochim. Biophys. Acra 1986,869,363. (d) Bobrowski, K.; Holcman, J.; Wierzchowski, K. L. Free Radical Res. Commun. 1989,6, 235. (e) Weinstein, M.; Alfassi, Z. B.; DcFelippis, M. R.; Klapper, M. H.; Faraggi, M. Biochim. Biophys. Acra 1991, 1076, 173. ( 5 ) (a) Prince, R. C. Trends Biochem. Sci. 1988, 13, 286. (b) Prince, R. C.; George, G. N. Ibid. 1990, 15, 170. (6) Marcus, R. A.; Sutin, N. Biochim. Biophys. Acra 1985, 811, 265. (7) (a) Sutin, N.; Brunschwig, B. S.;Creutz, C.; W i d e r , J. R. Pure Appl. Chem. 1988, 60, 1817. (b) Sutin, N. Prog. Inorg. Chem. 1983, 30, 441. (8) Ramachandran, G. N.; Sasisekharan, V. Adu. Protein Chem. 1968,23, 283. (9) (a) 'H and ')C NMR investigation on conformation of the title pep-
tides: (I) cis-trans isomerization about X-Pro peptide bonds and (ii) rotation about C,C, bonds of Trp and Tyr. (b) Effect of temperature on CD spectra and conformation of -(Pro)"- bridge. (c) Conformational energy calculations with use of the AMBER force field. (d) Full account of results obtainedga* will be published elsewhere. (10) (a) Siders, P.; Cave, R. J.; Marcus, R. A. J . Chem. Phys. 1984,81, 5613. (b) Cave, R. J.; Siders, P.; Marcus, R. A. J . Phys. Chem. 1986, 90, 1436. (c) Helms, A.; Heiler, D.; McLendon, G. J. Am. Chem. Sm. 1991,113, 4325.
(1 1) (a) Larsson, S.J . Chem. Soc.,Faraday Trans. 2 1983,79,1375. (b) Larsson, S.Chem. Phys. 1990,148, 103. (12) Newton, M. D. Chem. Rev. 1991, 91, 767. (13) (a) Beratan, D.; Onuchic, J. N.; Hopfield, J. J. J. Chem. Phys. 1987, 86,4488. (b) Onuchic, J. N.; Beratan, D. N. J. Chem. Phys. 1990,92,722. (c) Beratan, D.; Betts, J. N.; Onuchic, J. N. Science 1991, 252, 1285. (14) %hated, K.; Holcman, J.; Hart, E. J. J. Phys. Chem. 1983,87,1951. (IS) Bensasson, R. V.; Land, E. J.; Truscott, T. G. Flash Photolysis and Pulse Radiolysis; Pergamon Press: Oxford 1983; p 106. (16) (a) Weiner, S. J.; Kollman, P. A.; Case, D. A,; Singh, U. C.; Ghio, C.; Alagona, G.; Profeta, S.;Weiner, P. J . Am. Chem. Soc. 1984, 106, 765. (b) Singh, U. C.; Singh, P.; Weiner, J. C.; Kollman, P. University of California, San Francisco, 1986. (17) Jovanovic, S. V.; Steenken, S.;Simic, M. G. J . Phys. Chem. 1991, 95, 684.
The Journal of Physical Chemistry, Vol. 96, No. 24, 1992 10043 (18) Single isomers about Pro-Pro bonds in 2-5 were not considered because of their very low population (S5%)9.*dand similar distribution of r,, distances found in trial calculations. (19) (a) Grathwohl, Ch.; Wiithrich, K. Biopolymers 1981,20, 2623. (b) For H-TrpPro-OH at pD 7.7 and 298 K the rate constant for cis-trans isomerization is ,k = 3.5 X IO4 s-l, for peptides with non-C-terminal Pro it is larger by a factor of about 20;'" assuming from ref 19a k- 2 k,, and AGrr= 80 kJ mol-' one obtains T-' = (kF1 + kt+): =7 X and 1.5 X 10- s I at 298 and 328 K, respectively. (20) (a) Semisotnov, G. V.; Zikherman, K. Kh.; Kasatkin, S.B.; Ptitsyn, 0. B. Biopolymers 1981, 20,2287. (b) Nikoforovich, G. V.; Vesterman, B. G.; Bettins, J.; Podins, L. J. Biomol. Srrucr. Dynam. 1987, 4, 1119. (21) Vesterman, B. G.; Bobrowski, K.; Betins, J.; Nikoforovich, G. V.; Wierzchowski, K. L. Biochim. Biophys. Acra 1991, 1079, 39. (22) The most general form of the function k = A exp(-@r)[B + cos(C Oe)] was probed and found that B (and C) 0 while D = I . (23) In terms of electron tunneling through a square potential barrier: @ = (4Z(h)(h1!0)~/~, where m is the mess of electron and Vothe height of the potential barrier, equal, in effect, to a vertical ionization energy to the lowest unoccupied orbital in the medium spanning the two redox centers.6 For the aromatic s stems the value of the fl is about 1.1 A-'and corresponds to V, = 1.1 eV. ! 6 To explain a fl value about 10 A-',V, should be 1 order of magnitude higher than the observed ionization energy of tyrosine (-5 eV). (24) (a) Isied, S.S.;Vassilian, A.; Wishart, J. F.; Creutz, C.; Schwarz, H. A.; Sutin, N. J. Am. Chem. Soc. 1988,110,635. (b) Vassilian, A.; Wishart, J. F.; van Hemelryck, B.; Schwarz, H. A.; Isied, S.S.J . Am. Chem. Soc. 1990, 112,7278. (c) Isied, S.S.;Ogawa, M. Y.;Wishart, J. F. Chem. Reu. 1992,
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+
92, 381-394. (25) DeFelippis, M. R.; Murthy, C. P.; Broitman, F.; Weinraub, D.; Faraggi, M.; Klapper, M. H. J . Phys. Chem. 1991, 95, 3416. (26) Coefficients and @. now obtained differ somewhat numerically from those reported previously:'* 0.77 and -0.25 A-',due to the difference
in (r(CB))Roused in calculations. (27) (a) Schanze, K. S.;Sauer, K. J . Am. Chem. Soc. 1988, 1 IO, 1180. (b) Schanze, K. S.;Cabana, L. A. J. Phys. Chem. 1990, 94, 2740. (28) Bobrowski, K.; Holcman, J.; Ciurak, M.; Wierzchowski, K. L. Int. J. Radiar. Biol., in press. (29) Nemethy, G.; Scheraga, H. A. J . Phys. Chem. 1962,66, 1773. (30) (a) Mattice, W. L.; Mandelkem, L. Biochemisrty 1970,9, 1049. (b) Tiffany, M. L.; Krimm, S.Biopolymers 1972, 11, 2309. (c) Dukor, R. K.; Keiderling, T. A. Biopolymers 1991, 31, 1747. (31) About half of this shift is due to intrinsic thermodynamicsof cis-trans eq~ilibrium'~'and the other part arises from a large shift in the zwitterion anion equilibrium toward anionic form of the peptides (pK. of N-terminal Trp at 298 K of about 7.8 and 7.5 for trans and cis isomers, respectively; at 328 K it is lower by about 0.6 nit)^'.^^ coupled with a small shift of cis trans equilibrium toward the cis form." (32) Chang, M. C.; Petrich, J. W.; McDonald, D. B.; Fleming, G. R. J. Am. Chem. Soc. 1983,105,3819. (33) The apparent lack of this effect on AH' in other Pro-bridged systems (Os(II)/Ru(III)24 and d r ( R e ) / ~ * ( b p y ) ~is~ )most probably due to much higher rates of LRET therein, so that 6ka/6(r(T))
