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1381 Electron transport via metalloporphyrins EUGENE C. JOHNSON, TONY NIEM,l AND DAVID DOLPHIN 2 The Department ,!(Chelllistry. The /Jnil'ersitv oIBrilish Columbia. Valleoul'(!/", B.C.. Canada V6T I WS Received September 9, 1977 EUGENE C. JOHNSON, TONY NJEM, and DAVID DOLPHIN. Can. J. Chern. 56.1381 (1978). The controlled potential electrolysis of Ni(lI) lIleso-tetraphenylporphyrin (Ni(II)TPP) gives at room temperature the corresponding metalloporphyrin it-cation radical [Ni(II)TPP]+·. Upon freezing a solution of the it-cation radical to 77 K an internal electron transfer occurs to give [Ni(III)TPP] +. A discussion of the routes of electron transport in heme proteins is given, and the roles of metalloporphyrin it-cation radicals in electron transport is evaluated. EUGENE C. JOHNSON, TONY NIEM et DAVID DOLPHIN. Can. J. Chern. 56. 1381 (1978). L'electrolyse a potentiel contra Ie de la me.l'O-tetraphenylporphyrine de Ni(II) (Ni(lI)TPP) conduit, a temperature ambiante, au radical IT cation metalloporphyrine correspondant [Ni(Il)TPP)+·. Lorsque I'on congele une solution du radical IT cation it 77 K, un transfert interne d'electron se produit pour conduire au [Ni(lll)TPP] +. On discute des diverses routes de transport d'electron dans des proteines contenant de l'heme et on cvalue Ie role des radicaux it cation metalloporphyrine dans Ie transport des electrons. [Traduit par Ie journal] Introduction Redox reactions play critical roles in many biochemical transformations (I), and amongst the coenzymes and cofactors involved the iron porphyrins (hemes) play many varied and crucial functions. Hemes not only transport and store oxygen via hemoglobin and myoglobin, but in the form of cytochromes P-450 activate oxygen which, as monooxygenases bring about numerous biochemical oxidations (2). Other heme enzymes such as ca talase and the peroxidases act as two electron reductants of hydrogen peroxide to hydroxide, and are concomitantly oxidized by two electrons to the so-called primary complexes of the enzymes. In the case of catalase the primary complex then acts as a two electron oxidant of hydrogen peroxide to oxygen, and is reduced to the resting ferrihemoprotein. Another group of heme-containing proteins, the cytochromes, function between Fe(II) ~ Fe(III) couples, and by linking centres of different redox potcntials couple the free energy change of the electron transport to covalent bond formation (e.g. oxidative phosphorylation (3)). An insight into the modes of electron transport of metalloporphyrins was gained when it was shown that the initial, photochemically promoted, electron transport in photosynthesis involves the loss of an electron from one of the filled n:-molecular orbitals of chlorophyll a to generate a chlorophyll (or bacteriochlorophyll) n:-cation radical (4-6). Stable TC-cation radicals of simpler metalloporphyrins had been re'Work carried out by T.N. as a postdoctoral fellow in (he Chemistry department, Harvard University. 2Author to whom correspondence should be addressed. ported earlier (7), and shortly after their occurrence in photosynthetic systems was elucidated it was found that similar l't-cation radicals were the catalytically active forms of both the catalases and peroxidases (8, 9), Thus when either catalase or peroxidase is oxidized by hydrogen peroxide one electron is removed from the ferric iron and one from the TC-cloud to give, in the primary complexes, an Fe(IY) TC-cation radical electronic configuration, However, the ground states of these primary complexes differ in that in onc the remaining unpaired electron is in an a 2u , and in the other in an a lll orbital. The corresponding 2 A 2u and 2 Al u ground states, as a result of their different electronic ground states, exhibit different optical and enzymic properties (8). A change in the ground state from 2 Al u to 2 A 2u can bc demonstrated with the n:-cation radical of cobaltic octacthylporphyrin (7). Thus in the presence of bromide ion the cobalt coordinates two ligands to give [Co(III)OEP]2+' 2Br- which exhibits a 2 A lu ground state. When the bromide ions are removed and replaced by perchlorate the ground state changes to that of 2 A 2u' [t is apparent that small changes in the environment about the metal (and at the porphyrin periphery) can bring about a change in the ground state, and this is consistent with calculations which suggest that the two ground states differ in energy by about 1500-2000 cm - 1 or 4.3-5.7 kcal mol- 1 (10). We have been explicit in describing the electronic configuration of an oxidized metalloporphyrin and one might ask if it is possible to describe these configurations as resulting entirely from a metal or a ligand oxidation. Thus a cobaltic n:-cation radical which has two electrons less than the Co(I1) species 1382 CAN J. CIIEM. VOL. 56. 1978 could formally be described as both a derivative of cobalt(IV) or a n-dication of cobalt(Il). One might be inclined to conclude that" each of these descriptions are extreme resonance forms of the same molecule and that the molecule should be described as some contribution of all of them. This is not, however, th::: case since the epr spectrum of [Co(lll)OEP]2+' shows that the unpaired electron spends less than 5% of its time on the metal (9). Since the unpaired electron is in a ligand based orbital it can be described as a n-cation radical of cobaJt(III). Similarly the oxidation of ferric porphyrins removes electrons from the metal, not the ligand, since the optical spectra (which are dominated by rr-rr* transitions of the ligand) remain essentially unchanged upon oxidation whereas a dramatic change in the optical spectra would be observed if the ligand were oxidized (11). The ground state electronic contiguration of a metalloporphyrin can, from optical and epr measurements, be accurately determined; and the el ectronic configuration adopted will depend upon a variety of factors including the metal, the porphyrin and its peripheral substituents, and the axial ligands. Even in the 'simplest' metalloporphyrins small changes will influence the ground state. Thus the electrochemical one-electron oxidation of ruthenium(II) complexes of meso-tetraphenylporphyrin (TPP) gave a n-cation of [Ru(II)TPP](CO)(py) (viz. [Ru(II)TPP] + . (CO)(py)) but caused oxidation of the metal with [Ru(II)TPP](pY)2 to give [Ru(III)TPP] +(PY)2 (12). Hence the su bstitution of the weakly backbonding ligand pyridine for the more strongly backbonding CO destabilized Ru(IJ) vs. Ru(III) and changed the electronic configuration of the one-electron oxidized species. Even though the electronic configuration of a metalloporphyrin can now be readily determined, the mechanism, or route, of electron transfer by which such systems undergo redox chemistry is still a subject of much speCUlation with respect to even simple metalloporphyrins, which may be used as models of heme proteins; while the mechanisms by which heme proteins themselves transport electrons still remain, in general, largely unexplained (1). As with other coordination complexes metalloporphyrins have been found to effect electron transfer via two well-characterized mechanisms; inner sphere (13), where electron transfer occurs through a bridging species between oxidant and reductant, and outer sphere, where a bridging species is absent and electron transfer must occur through the extended rr-systems of the oxidant and reductant (14). While inner sphere mechanisms may operate with some model meta\loporphyrins it is apparent, on spatial considerations alone, that for most heme proteins in general an inner sphere mechanism cannot obtain. Wolberg and Manassen (15) have reported that the electrochemical oxidation ofNi(II)TPP in benzonitrile initially gives a Ni(III)TPP+ species which decayed to a Ni(II)TPP+' species. The presence of the two complexes was suggested by epr. In contrast we have recently reported that a similar oxidation in CH 2 Ci? gives the [Ni(II)TPP]+ . species, which is in thermo~ dynamic equilibrium with a [Ni(III)TPP]+ species at 77 K, as shown by epr and electronic absorption spectra (16). We describe here a detailed .study of these systems and suggest evidence of possible modes for outer sphere mechanisms of electron transfer in metalloenzymes. Experimental Materials Dichloromethane (Fischer, reagent) was ref1uxed over calcium hydride and distilled and storcd over 4 A molecular sieves. Tetra-n-butylammonium hexaAuorophosphate (TBAH) was prepared by treating an acetone solution of tetra-n-butylammonium iodide and ammonium hexafluorophosphate with water. The precipitated product was twice recrystallized from ethanol-water and dried in vacuum for 10 h at 100'C. Tetra-Ilbutylammonium perchlorate (TBAP) (Eastman) was twice recrystallized from ethanol-water and dried under vacuum for 10 h at 70°C. Tetraphenylporphyrinato nickel(II) was prepared from chlorin-free TPP since Ni(II)TPP is sparingly soluble and difficult to separate from chlorin impurity. 61Ni(II)TPP was prepared similarly except that 6l Ni (997,; enriched, Oakridge, TN) was initially dissolved in concentrated Hel, evaporated to dryness, and the resultant 6lNiCl 2 used in place of Ni(OAc)2. M easlIrements Ultraviolet-visible absorption spectra were recorded on a Cary 17 recording spectrophotometer. Room temperature spectra were recoded in CH 2 C1 2 using 1.0 cm cells. Spectra at 77 K were me<lsured using a variation of the opal glass method (17). A Pyrex cell was made by flattening a tube (0.5 cm OD) until the path length was ~0.1 mm. A quantity of a methylene dichloride solution was transferred to the cell by syringe and the sample frozen, evacuated, and sealed. After warming the cell to room temperature a strip of thin Teflon tape was stretched over the flat face and the cell was then placed in a windowed quartz Dewar filled with liquid nitrogen and placed in the sample beam of the spectrometer. A piece of opal glass was placed in the reference beam and optical spectra were then recorded as usual. The epr measurements, at both room temperature and 77 K, were made on a Varian E-3 EPR spectrometer in degassed methylene dichloride solutions in evacuated quartz epr tubes (4 mm 00). Electrochemical measurements were made at a Pt electrode in CHzCl z using 0.1 lv[ TBAH or TBAP as supporting electrolyte. A Ag/AgCI couple was used as reference. Standard 3 electrode operational amplifier circuitry was utilized as described previously (6). Due to the sparing solubility of Ni(II)TPP, special care was taken to ensure that no solid material was present during electrochemical or epr measurements. Solutions were either filtered prior to use or, for coulometric measurements, accurately weighted samples of ~ 3 mg were stirred for 30 min in 30 ml of degassed CH zCI 2 prior to use to 1383 JOHNSON ET AL. ensure complete solution. Such solutions were then used without further treatment. Results solutions of the second electrolysis product gave, by coulometry, n = 0.92 ± 0.05, but the appearance of additional cycles indicated decomposition. Spectra Electrochemistry Figure 2 follows the changes in the absorption The shape of the cyclic voltammograms varied significantly according to the supporting electrolyte spectra of Ni(II)TPP during electrolysis at 1.24 V to used. Figure I shows cyclic voltammograms of give the green one-electron oxidation product. The Ni(II)TPP in CH 2 Cl 2 using TBAH and TBAP. In the appearance of isosbestic points in the spectra indica tes presence of TBAP the cyclic voltammogram, as pre- a clean conversion to the oxidized form. A compariviously reported (16), is consistent with two overlap- son of the optical spectrum of the green oxidation ping electrochemically reversible couples centered at product to that of the well characterized IT-cation 1.17 and 1.29 V, while the data for the TBAH system radical of Mg(II)OEP (Fig. 3) shows that the product display two well-defined electrochemically reversible is the 1t-cation radical [Ni(lI)TPPt· and that further couples centred at 1.07 and 1.39 V. Exhaustive elec- electrolysis generates the corresponding rr-dication trolysis at 1.24 V in either system produced a green [Ni(II)TPP]2+ (Fig. 3) which has an optical spectrum solution which gave a cyclic voltammogram identical similar to other metalloporphyrin rr-dications. This to the starting material. Controlled potential coul- dication is a powerful electrophile reacting with a ometry (at 1.24 V) gave n-values of 1.01 ± 0.05, variety of nucleophiles such as methanol, or water, consistent with a one-electron oxidation of the to give isoporphyrins (18). Figure 4 shows the optical spectrum of [Ni(II)Ni(II)TPP. The green solution was stable for several days when protected from the light and the atmo- TPP] + ·PF 6 - at room temperature and after freezing sphere, and upon electrolysis at 0.80 V the starting to 77 K. The dramatic spectral changes are accommaterial was quantitatively regenerated (n = I), such panied by a change from a green solution to an that the cycle was completely reversible. Further electrolysis of the green oxidation product at potentials anodic to the second couple gave a brown solution which decomposed over a period of a few hours. 15 Immediate cathodic electrolysis of freshly prepared '" Q ~ '" a '" f'2?5.1SO (I ~ 20 10 . FIG. 2. Optical absorption spectra of Ni(II)TPP (A), during Its electrochemical oxidation, in CH,CJ 2 , to the x-cation radical [Ni(II)TPP] + . (E). b J20 10 A Q ~ '"\ 05 1.4 1.2 1.0 FIG. Volts vs Ag/AgCI 1. Cyclic voltammogram of Ni(IJ)TPP in methylene dichloride using a Ag/AgCI couple as reference; (a) tetra-nbutylammonium perchlorate and (b) tetra-n-butylammonium hexafluorophosphate as supporting electrolytes at 0.1 M. \ \ j~ FIG. 3. Optical absorption spectra, in CH 2 Cl 2 , of the x-cation radical [Ni(II)TPP] +. (A), the rc-dication [Ni(II)TPP]H (E). C is the optical spectrum during oxidation of A to B showing isosbestic points. D is the rc-cation radical [Mg(II)OEP] +. 1384 CAN. 1. CHEM. VOL. 56. 1978 no color change is observed. Similarly, these same solutions show no change in their epr spectra when cooled to 77 K, and only the isotropic signal at g = 2.0041 is present. FIG. 4. Optical absorption spectra, in CH 2 CI 2 , of [Ni(II)TPP] +. PF6 - at room temperature (A), the same sample at 77 K showing spectrum of [Ni(III)TPPJ+PF6 - (B), and Ni(II)TPP at 77 K (e). orange-red frozen solid which has a strong band at 526 nm. The spectra of both Ni(II)TPP and [Ni(II)TPPt· when measured in CH 2 Cl 2 in the presence of both TBAH and TBAP at 77 K show a band centered at 654 nm (in the absence of TBAP or TBAH Ni(II)TPP does not exhibit this absorption). In addition Ni(II)TPP+' at room temperature exhibits a broad absorption at 668 which moves to 656 at 77 K. The optical spectra of methylene dichloride solutions of the n-cation radical [Ni(II)TPP] +. [PF 6 or C10 4 showed no major changes upon cooling to 77 K when small amounts of water were present, nor were any changes observed with the bromide salt [Ni(II)TPPt· Br-. r Electron Paramagnetic Resonance The epr of the one-electron oxidation product were recorded at room temperature and at 77 K. At room temperature the green solution in the presence of PF 6 -, C10 4 -, or Br- exhibited a single signal, g = 2.0041 with a peak-to-peak separation of 47.2 G, indicative of a n-cation radical. At 77 K the orangered solid showed signals at gJ. = 2.286 and gil = 2.086, consistent with a low spin d 7 (Ni(III)) electronic configuration and in good agreement with spectra reported by Busch and co-workers (19) and by Wolberg and Manassen (15) for Ni(III) complexes. The epr signal of the Ni(III) species slowly decayed with time, being replaced after ~ 24 h by a weak isotropic signal which also decayed with time. We have already noted above that upon cooling solutions of the PF 6 - or C10 4 - salts, in the presence of small amounts of water, or the pure bromide salt Discussion From the physical studies described above we can conclude that when Ni(II)TPP is electrolyzed in methylene dichloride (with either TBAH, or TBAP as supporting electrolyte) at 1.24 V it undergoes a clean, reversible, one-electron oxidation to give the corresponding n-cation radical [Ni(II)TPPt·. At room temperature this species shows all of the properties of other similar, and well characterized, metalloporphyrin n-cation radicals with a 2 A2l1 ground state (5). The stability and ease of preparation of this n-cation radical is indicated by the clean electrochemistry and solution stability over long time periods. In addition this n-cation radical behaves like other metalloporphyrin n-cation radicals in that it can quantitatively be reduced back to Ni(II)TPP, and can be made to undergo a further one-electron oxidation to the n-dication [Ni(II)TPP]2+. In all of these respects [Ni(II)TPPt· is no different from other porphyrin n-cation radicals, which have been reported on in great detail, except for its quite exceptional thermal behavior. All of the above data are consistent with an internal electron transfer occurring as the temperature is lowered, with the hole jumping from the periphery to the metal. These redox reactions can be summarized as follows: -e -e Ni(II)TPP ~ [Ni(II)TPP] +. ~[Ni([I)TPPF + +e+e- HLl [Ni(III)TPP] + The strongest electronic transitions in metalloporphyrins (and their oxidized and reduced derivatives) are n -> n* in nature with molecular extinction coefficients offrom 10000 to 1 000000 (20, 21). In such systems the weak d-d transitions of the metal are hidden. Except when charge transfer between metal and ligand occurs, the oxidation state of a specific metal makes only minor perturbation on the optical transitions of metalloporphyrins. The spectra of Ni(II)TPP and that of the orange frozen samples of the n-cation radical [Ni(II)TPPt· are quite similar (Fig. 4) which show that all of the bonding 1tmolecular orbitals are filled. Since the n-cation radical has lost an electron from its highest filled bonding n-MO, then upon cooling an electron must move from the metal to fill the n-system and generate Ni(III). The formation of a nickel(III) complex at low temperatures is confirmed by epr spectroscopy which shows two signals having g values comparable to 1385 JOHNSON ET AL. those of previously reported Ni(III) complexes (I 7, 22). When the same series of epr experiments were carried out using 61Ni (J = 3/2) no change in the epr spectra of either the cation radical or the Ni(lII) species was observed. However, 61 Ni has a small magnetic moment ( - 0.75) resulting in a small hyperfine interaction which is lost under the relatively broad signal. W olberg and Manassen (15) have reported that the initial one-electron oxidation product of Ni(II)TPP in benzonitrile/TBAP is the metal oxidized [Ni(III)TPPt, and that upon standing at room temperature solutions of [Ni(III)TPPt began to show a weak epr signal which was assigned to [Ni(II)TPP] +'. The epr signal which they assigned to [Ni(III)TPPt was only observed at 77 K, and it is possible that these authors were observing a similar phenomenon to that reported here. However, the cyclic voltammogram reported for Ni(II)TPP in benzonitrilejTBAP and that reported here using methylene dichloride/ TBAP exhibit features which are atypical of most metalloporphyrins (22-25). In the former case the cyclic voltammogram showed two overlapping successive one-electron oxidations which were separated by not more than 100 mY. In the latter case we can resolve the two peaks at 1.17 and 1.29 V which differ by 120 mY. In the numerous cases where it has been shown that the two oxidations occur at the porphyrin periphery (to give rr-cation and rr-dication) the difference between the two potentials is 290 ± 50 m V (23), and deviations from this value suggest that the metal, rather than the ring, is the redox reactive centre. When Ni(II)TPP is oxidized in CH 2 CI 2 /TBAH the two oxidation waves are separated by 320 mV consistant with two ring oxidations, suggesting that the oxidations described above, which were carried out in the presence of perchlorate, involve an initial oxidation of the metal. In benzonitrile the (Ni(I1I)TPP]+ is sufficiently stable to be observed at room temperature before it converts over to the [Ni(II)TPP]+' whereas we find that at the end of a bulk electrolysis in methylene dichloride any [Ni(III)TPP]+ ,if formed, has already converted to (Ni(II)TPP] +'. Similar internal electron transfers have been reported previously (19,26), and Barefield and Mocella (27) showed that a nickel Curtis-type macrocycle exhibited a similar temperature-dependent internal electron transfer, i.e. 1 ¢ 2. Although the total charge on these complexes is 3 + instead of the I + for the systems reported here, the same general physical behavior was noted. Barefield attributed this phenomenon to the phase change in going from solution to the solid state, and suggested that axial ligation might play an important role in the process. A similar situation obtains here, for when a frozen sample is in thermal equilibrium with its solution the solid is orange while the solution is green, and only when the green solution freezes does it change color. It is not yet clear what happens, as the phase changes, to bring about the internal electron transfer and stabilize the hole on the metal rather than the periphery. One might anticipate that the higher oxidation state of the metal would be stabilized by the axial coordination of a donor ligand. The situation is not as clear cut as this, however, since in the presence Qf bromide or water only the n-cation radical is observed even at 77 K. Moreover, the fact that we have found no other prophyrin rr-cation which exhibits this internal electron transfer (including the n-cation radical of Ni(II) octaethylporphyrin) suggests that the phenomenon must depend upon a number of closely balanced factors such that small perturbations can stabilize one or another of the two species undergoing the internal electron transfer. Nonetheless, the degree of axial ligation does playa role here since in addition to the intense band at 525 nm the nickel(IIl) complex shows a broad weak absorption around 650 nm. While the origin of this charge transfer band is unclear, a similar band also occurs in the Ni(lI) complex, but only in the presence of ligands which would axially coordinate, and the appearance of a similar band in the spectrum of the Ni(III) com plex suggests that at 77 Keven PF 6 - and CI0 4 - are coordinating to the metal. Other metaIIoporphyrins show similar internal electron transfers. Ferguson et al. (28) while studying octaethylporphyrinato Pb(II) (Pb(II)OEP) found two reversible oxidations on the cyclic voltammetric time scale (s) which they attributed to successive, and reversible, one-electron oxidations of the ring. The resulting rr-dication decays, in rigorously purified solvents, with a half-life of ~ 5 min to a Pb(IV) com plex (6), i.e. -e -c Pb(II)OEP~ [Pb(II)OEP] + • .;=..-'[Pb(II)OEP]2+ t [Pb(IV)OEP]2+ Having shown that a facile and reversible internal electron transfer can occur between a metal and the periphery of a coordinating porphyrin one can ask could such an internal transfer play any role in biochemical systems? Even though many cytochromes function via an Fe(II)-Fe(III) couple, and while it is 1386 CAN. J. CHEM. VOL. 56, 197X of cytochrome c to form a channel from the surface of the protein to the heme. Similar remote pathways with an electron tunneling through several Angstroms Reference of a hydrophobic protein channel have been proReagent posed for some non-heme iron proteins, the ferre1,3 X 1011 29 enQ doxins (46). Thorough articulations of these argu8 Fe(dipy),(CN), + 1. 9 X 10 35 ments can be found elsewhere (47), but it should be 8,0 X 10 6 35 Fe(CN)6 3 37 6,25 X 10 6 Azurin noted such a pathway does not appear to be opera32 3,8 X 104 Ru(NH 3)62+ tive in cytochrome c. 2,6 X 10451 Fe(CN)6 4 (ii) A direct electron transfer through space-In 2 X 10452 Fe(II) cyt c addition to the classical inner- and outer-sphere 1.2 X 10433 Cr(Il) 32 Co(phenh'+ 1. 5 x 10 3 modes of electron transfer the possibilities that direct electron transfer via quantum mechanical electron tunneling, or transfer to and transport through a solabundantly clear that the final oxidation states are vent (as a hydrated electron (e aq - ) in biochemical indeed ferric and ferrous, how is ferrous cytochrome systems) must be considered. Even in redox reactions with simple inorganic ions the intermediacy, or lack oxidized to its ferric state? Much kinetic data have been gathered on the redox thereof, of eaq - has yet to be satisfactorily demonreactions of cytochromes, particularly the cyto- strated (48). The same situation applies to the more chromes c, using a variety of redox reagents ranging complex biochemical systems even though the reducfrom hydrated electrons (e aq - ) (29-31) through si m- tion of ferricytochrome c by externally generated pie inner- and outer-sphere inorganic reagents (32- eaq - has been reported (49). 36) to those macromolecules which transfer electrons Quantum mechanical electron tunneling is, howto and from cytochrome c in nature (37-41), Table I ever, well documented, and tunneling through dislists some of these redox reagents and observed rates tances of tens of Angstroms of inert solvents has of electron transfer, and while the rates vary they are been observed with non-biochemical systems (49). generally in the range of 10 3 -10 6 M - 1 S - 1. Cyto- While application of the simple one-dimensional chrome c contains an axially coordinated methionine theory (49) to complex macromolecules is unwar(42-43). Sutin has measured the rate of breaking the ranted, it does suggest that such mechanisms are Fe-S bond, and of opening a heme crevice, which possible in nature even though none has yet been should be rate limiting for an inner sphere pathway documented. (44). The measured value of 60 s -1 would appear to (iii) Oxidation at the porphyrin periphery-A numrule out an inner sphere process as a major route of ber of both kinetic and product ISO lation studies sugelectron transfer even with small inorganic redox gest that the oxidation of a coordinated metal can reagents. proceed through an outer-sphere mechanism, and X-ray crystallography has shown that both the studies with ferrocytochrome c and complexes of reduced and oxidized forms of cytochrome c have tris(i,lO-phenantholine)cobalt(IIl) ions show that an essentially the same structure (43). One striking feaouter sphere mechanism is operative and these studies ture of the crystal structure is the partially exposed have led Gray and co-workers (14) to suggest that edge of the heme porphyrin ring (a similar structural the point of electron transfer is the partially exposed feature is observed with cytochrome b s (45), and we heme edge. But having concluded that electron transanticipate that other cytochromes in the respiratory fer occurs via the porphyrin TC-cloud how, in fact, is chain will show similar exposed hemes). The tertiary the ferrocytochrome c oxidized to the ferric state? We structure of the protein will, of course, impose severe suggest that a process analogous to the oxidation of steric restraints upon the accessibility of the heme the Ni(II)TPP to the Ni(IIl)TPP+ might be operacoordinated iron to even the smallest redox reagent, tive, that is via the intermediacy of a porphyrin such that it is inconceivable that the iron atoms of TC-cation radical, such that the oxidation of ferro- to two adjacent cytochromes could come close enough ferricytochrome c could be descri bed as: to transfer an electron via an inner sphere mechanism. -eHow then do cytochromes transfer electrons amongst Fe(II) cyt c ~[Fe(II) cyt c]+';;=:=::':[Fe(lII) cyt c]+ themselves? Kinetic studies have suggested three +e main routes. III II (i) A remote transfer through aromatic residues in Since ferric and ferrous iron have different covalent the protein-which can be seen in the crystal structure TABLE I, Electron transfer rates for oxidation/reduction of cytochrome c JOHNSON ET AL. radii there must be conformational changes around the iron and its ligands between ferro- and ferricytochrome c (I and III) and between the postulated ferrocytochrome en-cation radical (II) and ferricytochrome c (III). Thus II and III would not be extreme resonance forms of the same species and an activation energy will be required to convert II to III. It has been suggested that the slow interconversion of [Pb(II)OEP]2+ to [Pb(IV)OEPf+, discussed above, may be due to a Frank-Condon forbidden transition since the Pb(II) is out of plane and the Pb(IV) in plane (50); and that since there is no overlap between the empty a 1u porphyrin orbital and the a2u(6p.) lead orbital the probability for charge transfer is small. 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