Spectroscopy and Molecular Modeling of Electrochemically Active

Downloaded via YORK UNIV on December 23, 2018 at 07:48:46 (UTC). See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.

Chapter 16

Spectroscopy and Molecular Modeling of Electrochemically Active Films of Myoglobin and Didodecyldimethylammonium Bromide 1

1

2

James T. Rusting , Alaa-Eldin F. Nassar , and Thomas F. Kumosinski 1

Department of Chemistry, P.O. Box U-60, University of Connecticut, Storrs, CT 06269-3060 Eastern Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, 600 East Mermaid Lane, Philadelphia, PA 19118 2

Water-insoluble coatings of didodecyldimethylammonium bromide (DDAB) on solid supports incorporate the protein myoglobin from solutions at p H 5.5-7.5 to form stable Mb -DDAB films. We previously found that electron transfer involving the heme Fe(III)/Fe(II) redox couple in 20 μm thick Mb-DDAB films on electrodes was 1000-fold faster than for Mb in aqueous solutions. The present work examines the supramolecular structure of Mb-DDAB films by reflectance -absorbance infrared, visible linear dichroism, and electron spin resonance spectroscopies. Molecular dynamics of Mb -DDAB models provided information on hydrophobic and coulombic interactions between Mb and DDAB. When combined with earlier thermal and electron transfer studies, results suggest that Mb-DDAB films feature lamellar liquid crystal DDAB arranged in bilayers with tilted hydrocarbon tails as in biological membranes. Mb in DDAB films has a secondary structure close to its native state, attains a preferred orientation in the films, and has Fe(III)heme in the high spin state. Mb electron transfer may be enhanced by adsorbed surfactant on the electrode which inhibits macromolecular impurities from adsorbing and blocking interfacial charge transfer. Biomembranes in living organisms are typically about half protein and half lipid. Many proteins and enzymes fulfill their biological functions as integral parts of membranes. The lipids are arranged in bilayers. Proteins can reside on the surface of, imbedded partly within, or extending across 0097-6156/94/0576-0250$08.00/0 © 1994 American Chemical Society Kumosinski and Liebman; Molecular Modeling ACS Symposium Series; American Chemical Society: Washington, DC, 1994.

16.

RUSLING ET AL.

Spectroscopy of ElectrochemicaUy Active Films

251

these bilayers (1). In this paper we discuss films of the protein myoglobin (Mb) and a surfactant which forms lamellar bilayer structures, didodecyldimethylammonium bromide (DDAB) (2). Myoglobin is a small oxygen carrying muscle protein with M W ca. 17,000. It contains a ferriheme prosthetic group that can be reduced electrochemically (3). Mb inserts spontaneously and rapidly from solution into cast DDAB films to form films stable for a month or more in buffers containing 50 m M NaBr (2). These films are in a lamellar liquid crystal phase at room temperature. When prepared on electrodes, the Fe(III) heme in Mb-DDAB films in the liquid crystal phase can be converted to Fe(II) heme at rates 1000-fold larger than for Mb in solution (2). Such films feature stacks of surfactant bilayers similar to lipid bilayers, and might provide useful experimental models for membrane-bound proteins. Our specific interest in Mb-DDAB derives from the desire to develop catalytic films to reductively dehalogenate organohalide pollutants at electrodes. In previous work, catalytic films were prepared from water insoluble cationic surfactants and negatively charged metal macrocyclic complexes such as phthalocyanine tetrasulfonates or corrin hexacarboxylates (4). DDAB is particularly suited for such applications since its films are in the liquid crystal phase at room temperature, facilitating efficient mass and charge transport necessary for catalytic applications. Mb reduces organohalides in its Fe(H) form in solution (5) and in DDAB films (2). Thus, Mb-DDAB films might serve as the basis for practical systems to dehalogenate or detect organohalide pollutants. They also might provide a model for reductive dehalogenation of pollutants in mammalian livers (6a) and anaerobic bacteria (6b). Kunitake et al. incorporated Mb into multibilayer surfactant films (7) prior to our work. The protein was reported to achieve a specific orientation in cast films of a double chain surfactant with an anionic phosphate head group (7). Possible explanations for the remarkable increase of electron transfer rate for Mb in DDAB films include (2) (i) the influence of strongly adsorbed surfactant in protecting the electrode from adsorption of macromolecules which can block electron transfer, and (ii) preferential orientation of Mb in a way favorable for electron transfer. Such orientation effects have been claimed for electrode surfaces chemically tailored to promote fast electron transfer to redox proteins (#). However, we are aware of very little direct structural evidence to support such views. In this paper, we describe structural studies on Mb-DDAB films combining ESR, UV-VIS linear dichroism, and infrared reflectanceabsorbance spectroscopy with molecular modeling and dynamics. The specific aim of this work is to characterize the supramolecular structure of the films. Such studies also provide insight into the influence of film structure on electron transfer kinetics of Mb in the film.

Kumosinski and Liebman; Molecular Modeling ACS Symposium Series; American Chemical Society: Washington, DC, 1994.

252

MOLECULAR MODELING

Experimental Section Materials. Lyophilized myoglobin (Mb) from horse skeletal muscle was from Sigma. Buffered myoglobin solutions were filtered through a YM30 filter (Amicon, 30,000 cutoff) to remove high molecular weight impurities (2). Tns-hydroxymethylaminomethane»HCl was used for p H 7.5 and acetate for p H 5.45 buffers, respectively. Buffers were 0.01 M in the conjugate base and contained 50 mM NaBr. Didodecyldimethylammonium bromide (DDAB) (>99%) was from Eastman Kodak. Water was purified with a Barnstead Nanopure system to a specific resistance >15 Megohm-cm. All other chemicals were ACS reagent grade. Apparatus and procedures. Cast DDAB films were prepared on solid substrates appropriate for each type of experiment. Briefly, a volume of 0.10.01 M DDAB in chloroform measured to give film thicknesses of 2-40 μπι, as necessary for a given experiment, was placed on the solid substrate with a microsyringe and spread evenly. Chloroform was evaporated gradually overnight (2). The cast films were then equilibrated in buffer solutions of 0.1-0.5 m M Mb for several hours. Uptake of Mb into the films monitored by cyclic voltammetry (CV) showed that steady state concentrations of Mb are achieved in 0°>45°. However, in the Mb-DDAB spectra, the signal to noise ratio at 100 Κ is not sufficient for quantitative measurement of the smaller peak. Although the data suggest orientation of Mb in the DDAB films, reliable estimates of orientation angles cannot be made. Electronic Spectra and Linear Dichroism. Absorbance of visible light by the ferriheme in Mb is responsible for a strong band around 410 nm, called the Soret band. A smaller band from the aromatic peptide side chains is found at 280 nm (Figure 4). The complexing strength of the labile axial ligand governs the position of the Soret band (14). Spectra of Mb in pH 5.5 and 7.5 buffers had the Soret band at 409 nm. Mb denatured with urea gave a band at 400 nm in buffer solutions, and at 405 nm in a DDAB film. Films cast from Mb alone gave Soret bands at 410 nm at p H 5.5 and 414 nm at p H 7.5, respectively. Values of λπ^χ in the MbDDAB film were shifted to 413 and 415 nm at p H 5.5 and 7.5, respectively (2). These values are closer to the 411 nm for crystalline high spin aquoMb than to those of 426 nm for CNMb crystals and 422 nm for azideMb crystals, both of which have strong axial ligands and are low spin (14). Thus, in agreement with IR and ESR data, electronic absorption spectra of MbDDAB films suggests that Mb is in its native, high spin form. Spectra had different intensities when obtained with parallel or perpendicularly polarized light (Figure 4). This observed linear dichroism was used to find an average order parameter S = (1-3 cos φ ) / 2 by using a simplified expression (15a) : 2

ΔΑ/Α = 3 S

(1)

where ΔΑ = A | |-Aj_ is the difference between the peak absorbances with parallel and perpendicularly polarized light, A=(A| ι + 2 A j _ ) / 3 is the absorbance of a randomly organized sample, and φ is the angle between the transition moment for the absorption and the normal to the film plane. Thus, the order parameter is found from: 5 = (Α,,-Αι)/(Α,,+2Αι)

(2)

Dichroic ratios A | ι /Aj_ for the Soret band as well as order parameters were significantly larger for Mb-DDAB films than for Mb films (Table II). These results suggest preferred average orientation for Mb in the DDAB films in agreement with conclusions from ESR. Since the transition


258

MOLECULAR MODELING

Table II. Linear Dichroic Ratios and Order Parameters from Soret Bands Mb films Mb-DDAB films pH* sample no. An/A| A||/Aj_ S" 7.5

1 2 3 4 5

1.32 1.16 1.16 1.27

0.097 0.051 0.050 0.085

7.5

(avg.± s.d.)

1.23±0.08

0.071±0.024

1.51 1.59 1.53 1.75 1.80 1.64±0.13

0.145 0.165 0.151 0.200 0.211 0.17±0.03

c

0.110 1.37 0.103 1.34 0.116 1.39 0.118 1.40 1.59 0.166 0.168 1.60 1.4510.12 0.13±0.03 5.5 1.14±0.03 0.045±0.009 (avs± s.d.) Buffers contained 50 mM NaBr. Order parameter from eq 2. Average values of φ for Mb-DDAB were 62±2 at p H 7.5 and 61±2° at p H 5.5, respectively. These values are not corrected for optical errors which could lead to bias on the order of 10% (15b). 5.5

1 2 3 4 5 6

1.11 1.14 1.13 1.13 1.19

0.036 0.045 0.043 0.040 0.060

c

a

b

c

moment of heme is in the plane of the molecule (14), φ represents the angle between the heme plane and the normal to the film plane. Values of φ in Table II (footnote c) are only approximate. More sophisticated experiments which correct for certain optical errors (15b) are underway. Molecular Modeling and Dynamics. The aim of this part of the work is to assess the theoretical viability of supramolecular models for Mb-DDAB films. Molecular models for bilayer films of DDAB were stable in the classic bilayer form with hydrocarbon tails together and charged head groups facing outward. The major fluctuations during dynamics were formation and disappearance of kinks and bends in the hydrocarbon tails. This is reminiscent of the mechanism for fluidity of the liquid crystal phase of bilayer membranes (1). Models of Mb alone featured many charge-paired partners of amino acid side chains on the globular protein surface. These charge pairs are reported (16) to impart stability to the secondary structure of Mb in water between p H 5 and 9. The globular structure of Mb also contains about 75% helix (17). A model of Mb-DDAB films was constructed from a bilayer of 48 DDABs surrounding a molecule of Mb. The model of Mb with unprotonated histidine residues (Color Plate 13) is roughly equivalent to its solution structure at p H 7.5. Side views of this model show that Mb extends about 10 À beyond the head group planes on both sides of the DDAB bilayer. When this model was allowed to undergo dynamics for 40 NOTE: The color plates can be found in a color section in the center of this volume.


16.

RUSLING ET AL.

Spectroscopy of Electrochemically Active Films

259

ps, its essential stability was evident. Mb stayed within the cavity in the DDAB bilayer and only a sort of breathing motion was observed around an average Mb-DDAB structure. Equilibrium was achieved within 40 ps, as shown by a time invariant radius of gyration. The charge paired amino acid partners of Mb can also be seen as pairs of red (negative) and purple (positive) amino acid residues close together on the Mb surface (Color Plate 13). Green hydrophobic residues remain within the interior, as with native Mb in water. This indicates that no large differences in secondary structure exist between Mb in water and Mb in the DDAB bilayer model. Similar results were found when the Mb histidine residues were protonated (Color Plate 14), representing the structure at p H 5.5. Again, many charge pairs were found on the Mb surface. This Mb-DDAB model was also stable during 40 ps dynamics runs. Since it is difficult to assess interactions of individual DDAB head groups with surface amino acid residues on Mb from the above models, a simpler model was constructed for this purpose. Two bilayer structures of four DDAB molecules each were docked with one set of head groups close to carboxylate side chains on the Mb surface (Figure 5a). During 40 ps dynamics computations, the surfactant head groups closest to Mb moved slightly away from the docking position, and the hydrocarbon chains of the DDABs waved about somewhat (Figure 5b). Furthermore, one of the bromide counterions moved well away from the protein surface. This suggests a weak electrostatic interaction between the head groups and the protein surface. A second set of similar models were constructed, but with the two D D A B mini-bilayers arranged with both sets of head groups and hydrocarbon tails close to the Mb surface (Figure 6a). This structure was also unstable. One set of hydrocarbon tails and one of the bromide ions moved away from the protein surface in 40 ps of dynamics (Figure 6b), although the DDAB clusters remained together. Discussion Film Structure and Stability. We first review previous findings on MbDDAB films (2), then integrate these with the results in this paper. Gel-toliquid crystal phase transition temperatures (T ) of Mb-DDAB films were 12 °C at p H 5.45 and 15 °C at pH 7.5. T was 15 °C for bilayer vesicles of DDAB, and 11 ° C for pure lamellar DDAB films. These T values similar to those of known DDAB bilayer systems suggest that DDAB in the films has a lamellar bilayer structure when Mb is present, and confirm that the films are liquid crystalline at ambient temperature. Our film thicknesses in the μιη range would represent thousands of bilayers in an ideal stacked lamellar film (4a). Scanning electron microscopy of freeze fractured cross sections of DDAB films suggest that these layered structures are somewhat wavy (4c), and contain significant defects. A wavy multilammelar structure has also c

c

c


MOLECULAR MODELING

260

1 cm s"). However, they are quite respectable c

6

_1

1


264

MOLECULAR MODELING

Table III. Summary of Electrochemical Parameters for Myoglobin at 25 ° C ref. pH sample lO^Dct, electrode cm /s V/NHE cm/s 2

5.5a 7.5 5.5a 5.5a 5.5-7.5a 7.0 a

Mb-DDAB-PG Mb-DDAB-PG Mb-CTAB-PGb Mb-SDS-PGb Η2θ, Mb/bare PG H2O, Mb/bare

0.54 0.41 0.55 0.30

7±1 8±1 3±2 2±1 ND 0.007

0.093 0.055 0.025 -0.030

d

—

—

0.5C

0.05

2 2 2 2 2 3

InSnU2 7.0 0.7 3 aq. CN-Mb/InSnO? -0.385 0.5C Solutions contained 50 m M NaBr. Results are averages from ref. 2 of values found by cyclic and normal pulse voltammetry. P G electrodes had adsorbed films of Mb and soluble surfactants sodium dodecylsulfate (SDS) and cetyltrimethylammonium bromide (CTAB). Diffusion coefficient of Mb in solution. Electron transfer not detected. a

b

c

d

for redox proteins, which often have difficulties exchanging electrons directly with electrodes (8). PG electrodes with coatings of adsorbed water soluble surfactants sodium dodecylsulfate (SDS) and cetyltrimethylammonium bromide (CTAB) also provide relatively fast electron transfer (Table ΙΠ). Comparison of standard potentials in the p H 7.5 film and in water at p H 7 (Table ΙΠ) suggests that the labile axial ligand on the Mb Fe(III)heme may be water. Charge transfer diffusion coefficients (D t) are also listed in Table ΙΠ. These are measures of the rate of transport of charge through the film when electrons are injected into it from the electrode. This is usually treated as a diffusion process (21). Values of D tfor Mb-DDAB are just slightly smaller than those of DDAB films loaded with ferrocyanide, cobalt(III)corrin hexacarboxylate, or metal phthalocyanine tetrasulfonates (4) which gave 0.6-1 χ 10 cm s for liquid crystal films. D t for Mb-DDAB in the gel state is 10-20 fold smaller (2). Films of ionic polymers loaded with electroactive counter ions are conceptually similar to DDAB films. D t for Mb-DDAB films are significantly larger than values of 10" to 10~ c m s for typical ionic polymers (21). Thus, even with the relatively large Mb molecule present, charge transport through liquid crystal DDAB films is faster than through most ionic polymer films. A common question for electroactive films is whether D t corresponds to physical diffusion of the electroactive species or to electronexchange between active sites, so called "charge hopping" (21). Measured times for breakthrough of Mb across DDAB films were consistent with those predicted for a molecule with D t = 4 χ 10~ cm s" (cf. Table ΙΠ) to achieve a root mean square displacement equivalent to film thickness.This suggests that Mb diffuses physically within Mb-DDAB films. c

c

-6

2

_1

c

c

8

10

2

_1

c

7

2

1

c


16.

RUSLING ET AL.

Spectroscopy of Electrochemically Active Films

265

Likely reasons for enhanced electron transfer rates in Mb-DDAB include the following: (i) strongly adsorbed surfactant on the electrode may inhibit competitive adsorption of macromolecular impurities in solution (3,8) which might otherwise block electron transfer; and (ii) Mb may be oriented in the film in a way favorable for electron transfer, for example with the heme group close to the electrode. Similar rate enhancements found with films of different types of surfactants (Table ΙΠ) suggest that surfactant adsorption on the electrode must play a role. However, this does not prohibit an influence for Mb orientation. ESR and linear dichroism suggest orientation of Mb in the DDAB films. These data do not as yet provide detailed structural information on the orientation of the key Mb molecules closest to the electrode surface where the electron transfer events occur. If molecular orientation plays a critical role in electron transfer kinetics, Mb molecules would have to achieve preferred orientations dynamically at the electrode-film interface during the voltammetric measurement. The electrode is most likely covered by a strongly adsorbed surfactant bilayer (22), before as well as after Mb is introduced into the film. If Mb resides on the outer surface of a bilayer of DDAB at the PG-film interface, it would be at least the width of a DDAB bilayer (30 À) away from the electrode. Transfer of an electron across such a distance is slow (23). These considerations suggest that Mb might be able to pass through the fluid DDAB bilayers during the dynamic electrochemical experiments. Studies of diffusion through biomembranes suggest that passage of proteins through intact bilayers may be slow (2). However, biomembranes of many organisms exist close to their phase transition temperatures, and may even contain small solid-like regions of gel-state lipids (24). On the other hand, the Mb-DDAB transition (T ) to the fluid liquid crystal phase occurs more than 10 ° C below room temperature. Thus, Mb-DDAB films may be in a more highly fluid state than typical biomembranes. High fluidity may permit a small protein like Mb to pass through more easily than if the film were close to its T . Consequently, the solid-like gel state retards charge and mass transport significantly, while the liquid crystal state facilitates these processes (2). The dynamic nature of surfactant bilayers also needs to be considered. Out of plane bending leading to transient pores or defects in biomembranes is now rather well accepted (1). These defects become larger in electric fields similar to those generated in voltammetry and can lead to bilayer rupture. Whether or not such processes can assist charge transport in DDAB films remains speculative at this stage of our work. c

c

Summary and Conclusions Spectroscopic and molecular modeling results presented herein along with earlier thermal and electron transfer studies can be combined to give a structural picture of Mb-DDAB films. The following features are established: (i) lamellar liquid crystal DDAB arranged in bilayers with tilted


266

MOLECULAR MODELING

ΐ'Ίΐ/Ύΐηι. Figure 7. Conceptual models of several bilayers of static Mb-DDAB films: (a) Mb resides partly within bilayers; (b) Mb resides between bilayers. hydrocarbon tails as in biological membranes; (ii) Mb in a high spin state with preferred orientation in the films; (iii) electron transfer enhanced by adsorbed surfactant on the electrode; (iv) charge transport facilitated by the fluid liquid crystal phase; and (v) physical diffusion of Mb within the films. In studies of multibilayer films of double chain phosphate surfactants containing Mb, Kunitake et al. proposed that Mb is intercalated between surfactant bilayers (7). At present we can imagine two limiting static models for the Mb-DDAB films, one with Mb within bilayers, but interacting in some way with adjacent bilayer head groups, and one with Mb between bilayers (Figure 7). A n intermediate supramolecular structure between these two extremes is also possible. Both limiting models might increase the stability of the films by electrostatic interactions of the DDAB head groups and the Mb surface amino acid residues or buffer components bound to Mb. However, because Mb has a positive charge at the pHs employed, direct surface interactions would have to be stronger than existing charge pair interactions of amino acid residues on the Mb surface. There is as yet no definitive experimental evidence in favor of either limiting model concerning the site of Mb residence in DDAB films. However, it is difficult to explain the electron transfer and charge transport results by the between-bilayer model (Figure 7b), unless Mb diffusion through defects in the films plays a dominant role in these processes. Such defects would be expected to be retained in both gel and liquid crystal states of a given film. Thus, the large difference in charge transport rate (2) above and below T might not occur if Mb were transported only through defects. Models featuring the possibility of Mb entry into fluid bilayers at the electrode-film interface go further at present to explain electrochemical results. Nevertheless, further studies are needed to achieve a more complete picture of the supramolecular structure of Mb-DDAB films. c

Acknowledgments. This work was supported by U.S. PHS grant No. ES03154 from NIH awarded by the National Institute of Environmental Health Sciences. The authors are grateful to Dr. Veeradej Chynwat for assistance with ESR and linear dichroism.


16. RUSLING ET AL.

Spectroscopy of Electrochemicatty Active Films

267

References and Notes (1) Kotyk, Α.; Janacek, K.; Koryta, J. Biophysical Chemistry of Membrane Structure, Wiley: Chichester, U. K., 1988, pp. 54-73. (2) Rusling, J. F.; Nassar, A. F. J. Am. Chem. Soc., 1993, 115, 11891-11847. (3) King, B. C.; Hawkridge, F. M.; Hoffman, Β. M . J. Am. Chem. Soc. 1992, 10603-10608. (4) (a) Rusling, J. F.; Zhang, H . Langmuir 1991, 7, 1791-1796. (b) Rusling, J. F.; H u , N . ; Zhang, H . ; Howe, D.; Miaw, C.-L.; Couture, E. in Electrochemistry in Colloids and Dispersions, Mackay, R. A . and Texter, J.

(Eds.), V C H Publishers; Ν , Y, 1992, pp. 303-318. (c) Hu, N.; Howe, D. J.; Ahmadi, M . F.; Rusling, J. F. Anal Chem. 1992, 64, 3180-3186. (d) Zhang, H.; Rusling, J. F.Talanta, 1993, 40, 741-747. (e) Miaw, C.-L.; Hu, N.; Bobbitt, J. M.; Ma, Z.; Ahmadi, M. F.; Rusling, J. F. Langmuir 1993, 9, 315-322. (5) (a) Wade, R. S.; Castro, C. E. J. Am. Chem. Soc. 1973, 95, 231-234. (b) Bartnicki, E. W.; Belser, N . O.; Castro, C. E. Biochemistry, 1978, 17, 55825586. (6) (a) Pryor, W. A. in Pryor, W. A. (Ed.) "Free Radicals in Biology" Academic: New York, 1976, pp. 1-50. (b) Brown, J. F.; Bedard, D. L . ; Brennan, M .J.;Carnahan, J. C.; Feng, H.; Wagner, R. F. Science, 1987, 236, 709. (7) (a) Hamachi, I.; Noda, S.; Kunitake, T. J. Am. Chem. Soc. 1990, 112, 6744-6745. (b) Hamachi, I.; Honda, T.; Noda, S.; Kunitake, T. Chem. Lett. 1991, 1121-1124. (c) Hamachi, I.; Noda, S.; Kunitake, T. J. Am. Chem. Soc. 1991, 113, 9625-9630. (8) (a) Armstrong, F. Α.; Hill, H . A . O.; Walton, N . J. Accts. Chem. Res. 1988, 21, 407-413. (b) Armstrong, F. A. in Bioinorganic Chemistry, Structure and Bonding 72, Springer-Verlag, Berlin, 1990, pp. 137-221. (9) Suga, K.; Rusling, J. F. Langmuir 1993, 9, 3649-3655. (10) (a) Press, W. H . ; Flannery, B. P.; Teukolosky, S. Α.; Vettering, W. T. Numerical Recipies Cambridge Univ. Press: Cambridge, MA: 1988, pp. 301327. (b) Weiner, S.J.;Coleman, P. Α.; Nguyen, D. T.; Case, D. A. J. Comput. Chem. 1986, 7, 230. (c) Hildebrand, J. H . Proc. Nat. Acad. Sci. 1979, 76, 194.

(11) Kauppinen, J. K.; Moffatt, D.J.;Mantsch, H . H.; Cameron, D. G. Appl. Spec. 1981, 35, 271-276. (12) Rusling, J. F.; Kumosinski, T. F. Intell. Instr. & Computers,

1992

(July/Aug.) 139-145. (13) Konetani, T.; Schleyer, H . J. Biol. Chem. 1967, 242, 3926. (14) Eaton, W. Α.; Hochstrasser, R. M . J. Chem. Phys. 1968, 49, 985-995. (15) (a) Breton, J.; Michel-Villaz, M . ; Paillotin, G. Biochim. Biophys. Acta 1973, 314, 42-56. (b) Norden, B.; Lindblom, G.; Jonas, I. J. Phys. Chem. 1977, 81, 2086-2093. (16) (a) Friend, S. H.; Gurd, F. R. N. Biochemistry 1979, 18, 4620. (b) Shire, S. J; Hanania, G. I. H.; Gurd, F. R. N . Biochemistry 1974, 13, 29672980. (c) Friend, S. H.; Gurd, F. R. N. Biochemistry 1979, 18, 4612-4619. (17) Kumosinski, T. F.; Farrell, H . , in Kumosinski, T. F.; Liebman, M . N . (Eds.); Molecular Modeling, this volume.


268

MOLECULAR MODELING

(18) Antonioni, E,; Rossi-Bernardi, L.; Chinacone, E. (Eds),Methods in Enzymology, Vol. 76 , Academic: N . Y., 1981, pp. 552-559. (19) Antonioni, E , ; Brunori, M . Hemoglobin

and Myoglobin

in their

Reactions with Ligands, North Holland: Amsterdam, 1971. (20) Klein, M . L . , Plenary Lecture, 67th Colloid and Surface Science Symposium, Toronto, Canada, June, 1993. (21) Charge transport diffusion through electroactive films is discussed by Murray, R. W. in Bard, A. J. (Ed.) Electroanalytical Chemistry, Vol. 13, Marcel Dekker, Ν. Y. 1984, pp. 191-368. (22) (a) Head down bilayers of DDAB and CTAB have been inferred from surface enhanced Raman spectroscopy in cast films on silver. There is evidence that head down orientation of these surfactants and SDS on electrodes occurs over a wide potential range when adsorption occurs from a relatively concentrated solution. (b) Suga, K.; Bradley, M.; Rusling, J. F. Langmuir , 1993, 9, 3063-3066. (c) Rusling, J. F. in Bard, A . J., E d , Electroanalytical Chemistry, Vol. 19, 1994, Marcel Dekker: New York, pp. 188. (23) (a) Closs, G.L.; Miller, J.R. Science, 1988, 240, 440.; (b) Mayo, S. L.; Ellis, W. R.; Crutchley, R.J.;Gray, H. B. Science, 1986, 233, 948. 22b

22c

(24) (a) Lee, A . G . Biochim. Biophys. Acta 1977, 472, 237-281; 285-344. (b)

Nagel; J. F. Ann. Rev. Phys. Chem. 1980, 31, 157-159. R E C E I V E D December 20, 1993


Spectroscopy and Molecular Modeling of Electrochemically Active

Recommend Documents