ELECTRON TRANSFER REACTIONS OCCURRING UPON LASER FLASH PHOTOLYSIS OF PHOSPHOLIPID BILAYER SYSTEMS CONTAINING CHLOROPHYLL, BENZOQUINONE AND CYTOCHROME c‐I. NEGATIVELY‐CHARGED VESICLES*

Abstract

Abstract— In negatively‐charged lipid bilayer vesicles prepared in deionized water from egg phosphatidylcholine and 25 mol % of α‐eleostearic acid, and containing chlorophyll a, benzoquinone, and cytochrome c, primary electron transfer after a laser flash occurred principally from chlorophyll triplet to benzoquinone, and to a smaller extent from chlorophyll triplet to oxidized cytochrome c. Several secondary electron transfer reactions occurred subsequent to this. The most rapid of these was electron transfer from reduced cytochrome c, which was bound to the outer surface of the negatively‐charged vesicle, to chlorophyll cation radical (k= 3.9 times 103 s‐1). Subsequent to this, the cation radical was reduced by benzoquinone anion radical (k= 1.6 times 102 s‐1>) and bound oxidized cytochrome c was reduced by the remaining anion radical which was expelled into the aqueous phase by the negative charge on the vesicle surface. This latter reaction occurred at the membrane‐solution interface with an observed rate constant (k= 60 s‐1) two orders of magnitude smaller than cytochrome oxidation. Net reduced cytochrome c was produced in this process. The reduced cytochrome c was slowly reoxidized by benzoquinone (k= 17 s‐1) and the system was returned to its original state. When the vesicle system was made slightly basic by adding tris(hydroxymethyl)aminomethane, the rates of both the reverse electron transfer between chlorophyll cation radical and benzoquinone anion radical (k= 5 times 102 s‐1) and the oxidation of reduced cytochrome c by chlorophyll cation radical (k= 9.4 times 103 s‐1) were accelerated. The rate of reduction of oxidized cytochrome c by benzoquinone anion radical remained approximately the same.

Surface plasmon resonance studies of complex formation between cytochrome c and bovine cytochrome c oxidase incorporated into a supported planar lipid bilayer. I. Binding of cytochrome c to cardiolipin/phosphatidylcholine membranes in the absence of oxidase

“…As has recently been demonstrated and discussed in detail below, SPR is a highly sensitive method for the direct measurement of protein binding to (Salamon et al, 1994bSoulages et al, 1995) or incorporation into (Salamon et al, 1994a(Salamon et al, , 1996 a solid-supported lipid bilayer, and is capable of detecting structural changes involving mass redistribution occurring within a proteolipid membrane (Salamon et al, 1994a). In previous work from this laboratory, kinetic methods have been used to study electron transfer between cyt c and other proteins in a lipid vesicle system (Fang and Tollin, 1988;Zhao and Tollin, 1991), and electrochemical techniques have been applied to measuring direct electron transfer from cyt c to a gold electrode, the surface of which was modified with a planar lipid membrane Tollin, 1991, 1993). These studies have demonstrated the importance of electrostatics in the electron transfer processes and have provided evidence which indicates that insertion of cyt c into the lipid film must occur to allow electron transfer through the membrane from the aqueous phase to the electrode surface.…”

QUINONES AS MEDIATORS OF ELECTRON TRANSFER FROM MEMBRANE‐BOUND CHLOROPHYLL TO CYTOCHROME c IN THE AQUEOUS PHASE IN LIPID BILAYER VESICLES: SYNERGISTIC ENHANCEMENT OF CYTOCHROME c REDUCTION BY LIPOPHILIC/ HYDROPHILIC QUINONE PAIRS

“…In these systems, 13-20% of the 3C molecules resulted in protein reduction, and the halftimes for reverse electron transfer ranged from 0.7-1 ms. We have also found that a viologen analog (propylene diquat) can mediate electron transfer from 'C to ferrodoxin in vesicles (Senthilathipan and Tollin, 1989). We have reported previously (Fang and Tollin, 1988a) that benzoquinone (BQ) can be used as a mediator of electron transfer from laser flash-excited 3C in negativelycharged lipid bilayer vesicles to cyt c (ox) located in the outer aqueous phase. The negative surface charge in this system aided in the expulsion of the similarly charged quinone anion radical from the bilayer interior into the aqueous interface region where the cytochrome was located.…”

Synthesis of Amphipathic Porphyrins and Their Photoinduced Electron Transfer Reactions at the Lipid Bilayer‐water Interface

“…The 10% impurity in the fraction containing 1 consisted ofa porphynn bearing a free formyl group (u = 1062.8) Treatment of this mixture with NaBH, (74 mg in 2 mL 95% ethanol) gave two porphyrin components that were readily separated by semipreparative centrifugal TLC ( I mm silica rotor, CH,CIZ), yielding 1 (1 16 mg, 11%) as the first band and the unwanted porphyrin (10 mg) as the second band. ' H NMR (CDCI,) 6 Synthesis of2. A sample of 1 ( I 0 mg, 0.01 1 mmol) was dissolved in 5 mL trifluoroacetic acid in a 25 mL one-neck round bottom flask.…”

“…"'0 They observed that introducing negative charges o n vesicle surfaces can increase the interfacial electron transfer rate between aqueous cytochrome c and membrane-bound chlorophyll cation. 6 In the present study, we investigate the effects of electrostatic interactions o n interfacial electron transfer in the lipid bilayer-water system by changing the charges of porphyrins and electron acceptors, and varying the ionic strength of the aqueous solutions. These ionic effects are usually described in terms of Gouy-Chapman theory,",'* but previous studies in this laboratory indicate that more local effects predominate in electron transfer r e a~t i 0 n s .…”