Electrostatic interactions stabilizing ferredoxin electron transfer complexes. Disruption by “conservative” mutations.

V.M Coghlan,L.E Vickery

doi:10.1016/s0021-9258(19)50369-6

V.M Coghlan, L.E Vickery

Open Access

https://doi.org/10.1016/s0021-9258(19)50369-6

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Abstract

Mitochondrial ferredoxins mediate electron transfer from NADPH:ferredoxin oxidoreductase to cytochrome P450 enzymes. Previous studies on human ferredoxin, in which acidic residues were replaced with neutral amino acids, established that Asp-76 and Asp-79 are are important for binding to both reductase and P450 (Coghlan, V. M., and Vickery, L. E. (1991) J. Biol. Chem. 266, 18606-18612). Here we report that replacement of Asp----Glu at position 76 or 79, whereas maintaining negative charge at these positions also results in dramatic decreases in binding affinity for both electron transfer partners (5-100-fold, delta(delta G) approximately 1.0-2.8 kcal/mol). These results imply that the active electron transfer complexes in these systems are dominated by a stable form which requires specific pairwise electrostatic interactions of fixed geometry for recognition and binding. This mechanism contrasts with that proposed for other electron transfer systems (as exemplified by cytochrome c) in which electrostatic interactions are believed to function primarily in precollisional orientation leading to "encounter complexes" having multiple geometries of similar free energy.

Highlights

Structures of complexes formed by soluble reversibly associating electron transfer proteins are noytet available,but both the observed variation in binding affinity with changes in ionic strength and theoretical calculations of potential fields suggest that electrostatic interactions play an important role
Because ferredoxin functions as asingle electron carrier, six association-dissociation interactions with Fd-reductase and P45Oscc occur as it shuttles reducing equivalents in the course of the overall reaction [17]
The vertebrate ferredoxins characterized to dateare all small (13-14 kDa) highly acidic proteins (PI- 4-4.5), and ionic strength effects on ferredoxin-mediated reactions suggest that electrostatic interactions are important forbinding to both redox partners [18].Fig. 1B shows a conserved region of negatively charged residues that occurs at positions 68-86

Summary

Introduction

Structures of complexes formed by soluble reversibly associating electron transfer proteins are noytet available,but both the observed variation in binding affinity with changes in ionic strength and theoretical calculations of potential fields suggest that electrostatic interactions play an important role. Because ferredoxin functions as asingle electron carrier, six association-dissociation interactions with Fd-reductase and P45Oscc occur as it shuttles reducing equivalents in the course of the overall reaction [17].

Results

Conclusion