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Intra- and inter-molecular electron transfer in cytochrome c and myoglobin observed by the muon spin relaxation method
Physica B 289}290 (2000) 631}635
Intra- and inter-molecular electron transfer in cytochrome c and myoglobin observed by the muon spin relaxation method K. Nagamine *, F.L. Pratt , S. Ohira , I. Watanabe , K. Ishida , S.N. Nakamura , T. Matsuzaki Muon Science Laboratory, RIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan Meson Science Laboratory, Institute of Materials Structure Science, KEK Tsukuba, Ibaraki, 305-0801, Japan
Abstract Rapid electron motion in the proteins, cytochrome c and myoglobin has been studied using muon spin relaxation. Measurements between 5 and 300 K show that the inter-site di!usion rate for the topologically 1D motion along the polypeptide chain is only weakly dependent on temperature. Evidence for an increase in higher dimensional motion is seen around 200 K in cytochrome c, apparently re#ecting structural change. For myoglobin the variation with temperature is di!erent, re#ecting the di!erent protein dynamics of this molecule. 2000 Elsevier Science B.V. All rights reserved. Keywords: Rapid electron transfer in protein; Spin labelling of moving electron; Motional narrowing
Electron transfer processes in macromolecules such as proteins are an important part of many biological phenomena, such as the storage and consummation of energy, photo-synthesis, etc. A number of experimental investigations have been carried out to explore the electron transfer phenomena in proteins and related chemical compounds. One of the key methods used to date for measuring long-distance electron transfer is to measure the electron-transfer kinetics using a #ash-quench procedure, thus concentrating on the speci"c path of the electron transfer [1]. However, almost all of the existing information on the * Corresponding address. Muon Science Laboratory, RIKEN 2-1 Hirosawa, Wako, Saitama 951-0198, Japan Fax: #81-48462-4648. E-mail addresses: [email protected], nagamine@ postman.riken.go.jp (K. Nagamine).
electron transfer has been obtained by essentially macroscopic methods. In order to understand the details of the electron transport it is thus very important to use methods that provide information at a more microscopic level. Among various types of proteins, cytochrome c has attracted much attention, since it plays an essential role in the respiratory electron transport chain in mitochondria, occupying a position next to the "nal process of the cycle, in which it transfers electrons to the surrounding oxidase complex [2]. On the other hand, myoglobin is known to serve an important role in oxygen transport with a basic molecular structure similar to that of electrontransfer protein. In order to obtain microscopic information on the electron transfer in the macro-molecule, the muon spin relaxation (lSR) method o!ers a great potential. During the slowing-down process, the
0921-4526/00/$ - see front matter 2000 Elsevier Science B.V. All rights reserved. PII: S 0 9 2 1 - 4 5 2 6 ( 0 0 ) 0 0 2 9 8 - 2
632
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
injected l> picks up one electron to form a neutral atomic state called muonium, which is in many ways analogous to a hydrogen atom. The muonium is then thermalized followed by chemical bonding to a reactive site on the molecule. Then, depending upon the nature of the molecule, the electron brought in by the l> can take on several characteristic behaviours, including localization to form a radical state and/or a linear motion along the molecular chain. These behaviours, by setting a time-origin of electron movement, can be detected most sensitively by measuring the spin relaxation process of the l> using the lSR method, which occurs through a magnetic interaction between the l> and the moving electron produced by the l> itself. This idea of sensitive detection of the electron behaviour in macromolecules using muons has been applied successfully in studies of the electron transport in conducting polymers [3}5]. A solitonlike motion has been studied for the l>-produced electron in trans-polyacetylene, which contrasts with the localization seen in cis-polyacetylene following the formation of a radical state [3,4]. Similarly, polaron-type electron transport phenomena in polyaniline has also been studied [5]. In longitudinal relaxation measurements, due to the nature of the interaction between the moving electrons and the stationary muons, the characteristic dimensionality of the electron motion can be studied by the dependence of the muon spin relaxation rate (j ) upon the externally applied magnetic l "eld (B ); for one-dimensional electron motion j J(B )\ , for two-dimensional electron l motion k J (a!b log B ) and for three-dimenl sional electron motion k does not usually have l signi"cant B dependence [6]. Progress has been made in the theoretical understanding of this paramagnetic relaxation process by Risch and Kehr who considered the direct stochastic treatment of the random-walk process of a spin which is rapidly di!using along a topologically one-dimensional chain [7]. An error-function-type longitudinal relaxation function (hereafter called the R}K function) G(t)"exp(Ct) erfc(Ct) was proposed for jt 1, where j is the electron spin
#ip rate, t the experimental time scale and
C a relaxation parameter. In this theoretical treatment, C is proportional to 1/B .
As a natural extension of this type of application of the lSR method, possible studies of electron transfer in protein were proposed in 1987 by one of the present authors [8]. Experiments on the l> relaxation in cytochrome c and myoglobin have been conducted using an intense pulsed beam (70 ns pulses at 50 Hz repetition rate) of 4 MeV l> at the RIKEN-RAL Muon Facility at Rutherford Appleton Laboratory, UK. The cytochrome c used here is Fe(3#) type in a polycrystalline powder form extracted from horse heart (Wako-Chemical product), while myoglobin was extracted from horse muscle (Sigma Co. Ltd.). As references, cytochrome c with Fe(2#) prepared by Drs M. Ataka and T. Kubota and lysozyme extracted from chicken egg (Sigma Co. Ltd.) were used. The present l>SR measurements were carried out for a range of temperatures between 5 and 300 K and for a number of longitudinal magnetic "elds in the range 0}0.4 T. All measurements were conducted on the powder sample as received. At each of the measurement temperatures, the l> relaxation function was found to have an external "eld dependence in the cases of cytochrome c with Fe(3#) and myoglobin. Some typical relaxation curves are shown in Fig. 1; it can be seen that the spin relaxation becomes suppressed at higher external "elds B , while the initial asymmetry at t"0, a (0) increases against B . The observed rel laxation functions G(t) were "tted with the R}K function [7] and the longitudinal relaxation parameter C obtained at various temperatures is seen to decrease monotonically with increasing B (Fig. 2). On closer inspection of the B dependence of C, there are two components, (1) a region of weak "eld dependence (lower "eld) and (2) a (B )\ dependent region (higher "eld). The latter region exhibits the characteristic l> spin relaxation behaviour due to linear motion of a paramagnetic electron. As seen in Fig. 2, the critical "eld (hereafter we refer to this as the cuto! "eld) where the second region takes over from the "rst region has a signi"cant temperature dependence; the cuto! "eld is seen to reduce with decreasing temperature. Similar measurements done for both lysozyme and cytochrome c with Fe (II) did not show any similar region where C follows a (B )\ dependence.
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
633
Fig. 1. Typical l> spin relaxation time spectra in cytochorome c at 5, 110 and 280 K under external longitudinal "elds of 0, 50 and 500 G. For "nite "elds the curves show "ts using the R}K function.
All of these results lead to the following microscopic picture of the electron transfer process for the muon-injected electron in cytochrome c and myoglobin. 1. Topologically linear motion along the chain of the cytochrome c. 2. The inter-site di!usion rate along the chain D , which can be obtained from the measured C and the hyper"ne coupling constant (500 MHz) estimated from the recovery curve of the initial asymmetry, becomes a value of the order of 10 rad/s and almost temperature independent (Fig. 3). This value is encouragingly consistent with those obtained using optical methods in Ruthenated proteins for short electron transfer distances in the region of 5 As [9]. 1. The cuto! process represents departure on a longer time-scale from the topologically onedimensional di!usion which occurs on the shorter time-scale. A naive picture based upon the previous l>SR studies on high molecular weight conducting polymers [3}5] would suggest an increase in the e!ective dimensionality of the
di!usion at temperatures higher than 200 K due to an increased inter-chain di!usion rate D . , Assuming that the crossover occurs as the interchain di!usion rate becomes comparable with the electron Larmor frequency [5], one can obtain the inter-chain di!usion rate D . Depend, ing upon the temperature region, the obtained D in cytochrome c is dominated by two di!er, ent processes as seen in the temperature dependence (Fig. 3); one with a characteristic activation energy of 150 meV seen above 200 K and the other with an activation energy of less than 2 meV seen below 200 K. The characteristic change at 200 K of the D seems to be related to , the well-known structural change of some proteins which has been suggested to be a glass-like transition [10]. In the context of a protein such as cytochrome c with coils and folds in its structure, the &inter-chain' di!usion might perhaps be interpreted as &inter-loop' jumps, which could well be strongly activated by the increased thermal displacement of the polymer occurring above the glass transition. On the other hand, D in myoglobin shows only one component, ,
634
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
Fig. 2. The R}K relaxation parameter C versus external logitudinal magnetic "eld for the l> in cytochrome c at typical temperatures. The B\-dependent part can be seen to become signi"cant in the higher "eld region and the critical "eld (cuto! "eld) of the onset of the B\ dependence can be seen to have a clear temperature dependence.
re#ecting the di!erent protein dynamics of this molecule for inter-chain electron transfer. The most important unknown pieces of information are the distribution of locations of the
l> bonding-sites and the corresponding electronic structure around the l> at the site from where the electron starts its linear motion. For this purpose, muon spin RF resonance has been conducted on the l> in both cytochrome c and myoglobin. The
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
635
e$ciency of the measurements should be emphasized, has the ability to extend the studies to cytochrome c in various chemical and biological environments; e.g. in solution with di!erent values of pH, etc.
Acknowledgements We acknowledge helpful discussions with Professor N. Go, K. Kehr and T. P. Das and Drs. M. Ataka, T. Kubota, N. Igarashi, T. Iizuka and A. Kira. The authors would like to thank Professors S. Kobayashi, A. Arima, M. Oda and other related persons at RIKEN and Drs. P. Williams, A.D. Taylor and W.G. Williams and other related persons at RAL for their kind support and encouragement in the development of the RIKEN-RAL Muon Facility.
Fig. 3. The upper plot shows the temperature dependence of the parallel di!usion rate of an electron in cytochrome c and myoglobin derived from the B\-dependent part of the relaxation parameter. The lower plot shows the perpendicular di!usion rate, derived from the cuto! "eld determined in Fig. 2, plotted against inverse temperature.
results gave some 10 ppm paramagnetic shift with a characteristic temperature dependence; more precise measurements will provide an information of the distance of the l> location from the heme Fe. Although, as mentioned above, there are still some unknown factors at this stage concerning the detailed nature of the l> probe state, it is clear that the electron transfer process through a microscopic section of cytochrome c was directly detected in the present experiment. The present lSR method should be contrasted with the experiments using photo-excited electrons, where electron transfer was measured through a path connecting two speci"c sites [11]. The l>SR method, where the high
References [1] T.B. Karpishin, M.W. Grisnta!, S. Komer-Panicucci, G. Mclendon, H.B. Gray, Structure 2 (1994) 415. [2] R.A. Scott, A.G. Mauk, University Science Books, Sausalito, California, 1995. [3] K. Nagamine, K. Ishida, T. Matsuzaki, K. Nishiyama, Y. Kuno, T. Yamazaki, H. Shirakawa, Phys. Rev. Lett. 53 (1984) 1763. [4] K. Ishida, K. Nagamine, T. Matsuzaki, Y. Kuno, T. Yamazaki, E. Torikai, H. Shirakawa, J.H. Brewer, Phys. Rev. Lett. 55 (1985) 2209. [5] F.L. Pratt, S.J. Blundell, W. Hayes, K. Nagamine, K. Ishida, A.P. Monkman, Phys. Rev. Lett 79 (1997) 2855. [6] M.A. Butler, L.R. Walker, Z.G. Soos, J. Chem. Phys. 64 (1976) 3592. [7] R. Risch, K.W. Kehr, Phys. Rev. B 46 (1992) 5246. [8] K. Nagamine, Invited Talk at Gordon Conference on Rapid Electron Transfer in Macromolecules, Los Angeles, 1987. [9] C.C. Moser, J.M. Keske, K. Warncke, R.S. Farid, P.L. Dutton, Nature 355 (1992) 796. [10] F. Parak, E.N. Frolov, R.L. MoK ssbauer, V.I. Godanskii, J. Mol. Biol. 145 (1981) 825. [11] S.E. Peterson-Kennedy, J.L. McGourty, J.A. Kalweit, B.M. Ho!man, J. Am. Chem. Soc. 108 (1986) 1739.
Intra- and inter-molecular electron transfer in cytochrome c and myoglobin observed by the muon spin relaxation method K. Nagamine *, F.L. Pratt , S. Ohira , I. Watanabe , K. Ishida , S.N. Nakamura , T. Matsuzaki Muon Science Laboratory, RIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, Saitama, 351-0198, Japan Meson Science Laboratory, Institute of Materials Structure Science, KEK Tsukuba, Ibaraki, 305-0801, Japan
Abstract Rapid electron motion in the proteins, cytochrome c and myoglobin has been studied using muon spin relaxation. Measurements between 5 and 300 K show that the inter-site di!usion rate for the topologically 1D motion along the polypeptide chain is only weakly dependent on temperature. Evidence for an increase in higher dimensional motion is seen around 200 K in cytochrome c, apparently re#ecting structural change. For myoglobin the variation with temperature is di!erent, re#ecting the di!erent protein dynamics of this molecule. 2000 Elsevier Science B.V. All rights reserved. Keywords: Rapid electron transfer in protein; Spin labelling of moving electron; Motional narrowing
Electron transfer processes in macromolecules such as proteins are an important part of many biological phenomena, such as the storage and consummation of energy, photo-synthesis, etc. A number of experimental investigations have been carried out to explore the electron transfer phenomena in proteins and related chemical compounds. One of the key methods used to date for measuring long-distance electron transfer is to measure the electron-transfer kinetics using a #ash-quench procedure, thus concentrating on the speci"c path of the electron transfer [1]. However, almost all of the existing information on the * Corresponding address. Muon Science Laboratory, RIKEN 2-1 Hirosawa, Wako, Saitama 951-0198, Japan Fax: #81-48462-4648. E-mail addresses: [email protected], nagamine@ postman.riken.go.jp (K. Nagamine).
electron transfer has been obtained by essentially macroscopic methods. In order to understand the details of the electron transport it is thus very important to use methods that provide information at a more microscopic level. Among various types of proteins, cytochrome c has attracted much attention, since it plays an essential role in the respiratory electron transport chain in mitochondria, occupying a position next to the "nal process of the cycle, in which it transfers electrons to the surrounding oxidase complex [2]. On the other hand, myoglobin is known to serve an important role in oxygen transport with a basic molecular structure similar to that of electrontransfer protein. In order to obtain microscopic information on the electron transfer in the macro-molecule, the muon spin relaxation (lSR) method o!ers a great potential. During the slowing-down process, the
0921-4526/00/$ - see front matter 2000 Elsevier Science B.V. All rights reserved. PII: S 0 9 2 1 - 4 5 2 6 ( 0 0 ) 0 0 2 9 8 - 2
632
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
injected l> picks up one electron to form a neutral atomic state called muonium, which is in many ways analogous to a hydrogen atom. The muonium is then thermalized followed by chemical bonding to a reactive site on the molecule. Then, depending upon the nature of the molecule, the electron brought in by the l> can take on several characteristic behaviours, including localization to form a radical state and/or a linear motion along the molecular chain. These behaviours, by setting a time-origin of electron movement, can be detected most sensitively by measuring the spin relaxation process of the l> using the lSR method, which occurs through a magnetic interaction between the l> and the moving electron produced by the l> itself. This idea of sensitive detection of the electron behaviour in macromolecules using muons has been applied successfully in studies of the electron transport in conducting polymers [3}5]. A solitonlike motion has been studied for the l>-produced electron in trans-polyacetylene, which contrasts with the localization seen in cis-polyacetylene following the formation of a radical state [3,4]. Similarly, polaron-type electron transport phenomena in polyaniline has also been studied [5]. In longitudinal relaxation measurements, due to the nature of the interaction between the moving electrons and the stationary muons, the characteristic dimensionality of the electron motion can be studied by the dependence of the muon spin relaxation rate (j ) upon the externally applied magnetic l "eld (B ); for one-dimensional electron motion j J(B )\ , for two-dimensional electron l motion k J (a!b log B ) and for three-dimenl sional electron motion k does not usually have l signi"cant B dependence [6]. Progress has been made in the theoretical understanding of this paramagnetic relaxation process by Risch and Kehr who considered the direct stochastic treatment of the random-walk process of a spin which is rapidly di!using along a topologically one-dimensional chain [7]. An error-function-type longitudinal relaxation function (hereafter called the R}K function) G(t)"exp(Ct) erfc(Ct) was proposed for jt 1, where j is the electron spin
#ip rate, t the experimental time scale and
C a relaxation parameter. In this theoretical treatment, C is proportional to 1/B .
As a natural extension of this type of application of the lSR method, possible studies of electron transfer in protein were proposed in 1987 by one of the present authors [8]. Experiments on the l> relaxation in cytochrome c and myoglobin have been conducted using an intense pulsed beam (70 ns pulses at 50 Hz repetition rate) of 4 MeV l> at the RIKEN-RAL Muon Facility at Rutherford Appleton Laboratory, UK. The cytochrome c used here is Fe(3#) type in a polycrystalline powder form extracted from horse heart (Wako-Chemical product), while myoglobin was extracted from horse muscle (Sigma Co. Ltd.). As references, cytochrome c with Fe(2#) prepared by Drs M. Ataka and T. Kubota and lysozyme extracted from chicken egg (Sigma Co. Ltd.) were used. The present l>SR measurements were carried out for a range of temperatures between 5 and 300 K and for a number of longitudinal magnetic "elds in the range 0}0.4 T. All measurements were conducted on the powder sample as received. At each of the measurement temperatures, the l> relaxation function was found to have an external "eld dependence in the cases of cytochrome c with Fe(3#) and myoglobin. Some typical relaxation curves are shown in Fig. 1; it can be seen that the spin relaxation becomes suppressed at higher external "elds B , while the initial asymmetry at t"0, a (0) increases against B . The observed rel laxation functions G(t) were "tted with the R}K function [7] and the longitudinal relaxation parameter C obtained at various temperatures is seen to decrease monotonically with increasing B (Fig. 2). On closer inspection of the B dependence of C, there are two components, (1) a region of weak "eld dependence (lower "eld) and (2) a (B )\ dependent region (higher "eld). The latter region exhibits the characteristic l> spin relaxation behaviour due to linear motion of a paramagnetic electron. As seen in Fig. 2, the critical "eld (hereafter we refer to this as the cuto! "eld) where the second region takes over from the "rst region has a signi"cant temperature dependence; the cuto! "eld is seen to reduce with decreasing temperature. Similar measurements done for both lysozyme and cytochrome c with Fe (II) did not show any similar region where C follows a (B )\ dependence.
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
633
Fig. 1. Typical l> spin relaxation time spectra in cytochorome c at 5, 110 and 280 K under external longitudinal "elds of 0, 50 and 500 G. For "nite "elds the curves show "ts using the R}K function.
All of these results lead to the following microscopic picture of the electron transfer process for the muon-injected electron in cytochrome c and myoglobin. 1. Topologically linear motion along the chain of the cytochrome c. 2. The inter-site di!usion rate along the chain D , which can be obtained from the measured C and the hyper"ne coupling constant (500 MHz) estimated from the recovery curve of the initial asymmetry, becomes a value of the order of 10 rad/s and almost temperature independent (Fig. 3). This value is encouragingly consistent with those obtained using optical methods in Ruthenated proteins for short electron transfer distances in the region of 5 As [9]. 1. The cuto! process represents departure on a longer time-scale from the topologically onedimensional di!usion which occurs on the shorter time-scale. A naive picture based upon the previous l>SR studies on high molecular weight conducting polymers [3}5] would suggest an increase in the e!ective dimensionality of the
di!usion at temperatures higher than 200 K due to an increased inter-chain di!usion rate D . , Assuming that the crossover occurs as the interchain di!usion rate becomes comparable with the electron Larmor frequency [5], one can obtain the inter-chain di!usion rate D . Depend, ing upon the temperature region, the obtained D in cytochrome c is dominated by two di!er, ent processes as seen in the temperature dependence (Fig. 3); one with a characteristic activation energy of 150 meV seen above 200 K and the other with an activation energy of less than 2 meV seen below 200 K. The characteristic change at 200 K of the D seems to be related to , the well-known structural change of some proteins which has been suggested to be a glass-like transition [10]. In the context of a protein such as cytochrome c with coils and folds in its structure, the &inter-chain' di!usion might perhaps be interpreted as &inter-loop' jumps, which could well be strongly activated by the increased thermal displacement of the polymer occurring above the glass transition. On the other hand, D in myoglobin shows only one component, ,
634
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
Fig. 2. The R}K relaxation parameter C versus external logitudinal magnetic "eld for the l> in cytochrome c at typical temperatures. The B\-dependent part can be seen to become signi"cant in the higher "eld region and the critical "eld (cuto! "eld) of the onset of the B\ dependence can be seen to have a clear temperature dependence.
re#ecting the di!erent protein dynamics of this molecule for inter-chain electron transfer. The most important unknown pieces of information are the distribution of locations of the
l> bonding-sites and the corresponding electronic structure around the l> at the site from where the electron starts its linear motion. For this purpose, muon spin RF resonance has been conducted on the l> in both cytochrome c and myoglobin. The
K. Nagamine et al. / Physica B 289}290 (2000) 631}635
635
e$ciency of the measurements should be emphasized, has the ability to extend the studies to cytochrome c in various chemical and biological environments; e.g. in solution with di!erent values of pH, etc.
Acknowledgements We acknowledge helpful discussions with Professor N. Go, K. Kehr and T. P. Das and Drs. M. Ataka, T. Kubota, N. Igarashi, T. Iizuka and A. Kira. The authors would like to thank Professors S. Kobayashi, A. Arima, M. Oda and other related persons at RIKEN and Drs. P. Williams, A.D. Taylor and W.G. Williams and other related persons at RAL for their kind support and encouragement in the development of the RIKEN-RAL Muon Facility.
Fig. 3. The upper plot shows the temperature dependence of the parallel di!usion rate of an electron in cytochrome c and myoglobin derived from the B\-dependent part of the relaxation parameter. The lower plot shows the perpendicular di!usion rate, derived from the cuto! "eld determined in Fig. 2, plotted against inverse temperature.
results gave some 10 ppm paramagnetic shift with a characteristic temperature dependence; more precise measurements will provide an information of the distance of the l> location from the heme Fe. Although, as mentioned above, there are still some unknown factors at this stage concerning the detailed nature of the l> probe state, it is clear that the electron transfer process through a microscopic section of cytochrome c was directly detected in the present experiment. The present lSR method should be contrasted with the experiments using photo-excited electrons, where electron transfer was measured through a path connecting two speci"c sites [11]. The l>SR method, where the high
References [1] T.B. Karpishin, M.W. Grisnta!, S. Komer-Panicucci, G. Mclendon, H.B. Gray, Structure 2 (1994) 415. [2] R.A. Scott, A.G. Mauk, University Science Books, Sausalito, California, 1995. [3] K. Nagamine, K. Ishida, T. Matsuzaki, K. Nishiyama, Y. Kuno, T. Yamazaki, H. Shirakawa, Phys. Rev. Lett. 53 (1984) 1763. [4] K. Ishida, K. Nagamine, T. Matsuzaki, Y. Kuno, T. Yamazaki, E. Torikai, H. Shirakawa, J.H. Brewer, Phys. Rev. Lett. 55 (1985) 2209. [5] F.L. Pratt, S.J. Blundell, W. Hayes, K. Nagamine, K. Ishida, A.P. Monkman, Phys. Rev. Lett 79 (1997) 2855. [6] M.A. Butler, L.R. Walker, Z.G. Soos, J. Chem. Phys. 64 (1976) 3592. [7] R. Risch, K.W. Kehr, Phys. Rev. B 46 (1992) 5246. [8] K. Nagamine, Invited Talk at Gordon Conference on Rapid Electron Transfer in Macromolecules, Los Angeles, 1987. [9] C.C. Moser, J.M. Keske, K. Warncke, R.S. Farid, P.L. Dutton, Nature 355 (1992) 796. [10] F. Parak, E.N. Frolov, R.L. MoK ssbauer, V.I. Godanskii, J. Mol. Biol. 145 (1981) 825. [11] S.E. Peterson-Kennedy, J.L. McGourty, J.A. Kalweit, B.M. Ho!man, J. Am. Chem. Soc. 108 (1986) 1739.