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Roles of conserved Arg72 and Tyr71 in the ascorbate-specific transmembrane electron transfer catalyzed by Zea mays cytochrome b561

Paper ID Volume ID Publish Year Pages File Format Full-Text
20867 43195 2013 10 PDF Available
Title
Roles of conserved Arg72 and Tyr71 in the ascorbate-specific transmembrane electron transfer catalyzed by Zea mays cytochrome b561
Abstract

Cytochromes b561, novel transmembrane electron transport proteins residing in eukaryotic cells, have a number of common features including six transmembrane α-helices and two heme ligation sites. Our recent studies on recombinant Zea mays cytochrome b561 suggested that concerted proton/electron transfer mechanism was functioning in plant cytochromes b561 as well and that conserved Lys83 on a cytosolic loop had important roles for ascorbate-binding and a succeeding electron transfer. In the present study, we conducted site-directed mutagenesis analyses on conserved Arg72 and Tyr71. Removal of a positive charge at Arg72 did not affect significantly on the final heme reduction level with ascorbate as reductant. However, characteristic pH-dependent initial time-lag upon electron acceptance from ascorbate was completely lost for R72A and R72E mutants. Substitution of Tyr71 with Ala or Phe affected both on the final heme reduction level and on the pH-dependent initial time-lag, causing acceleration of the electron transfer. These observations were interpreted as existence of specific interactions of Tyr71 and Arg72 with ascorbate. However, their mechanistic roles were distinctly different from that of Lys83, as exemplified by K83A/Y71A double mutant, and might be related for expelling of monodehydroascorbate radical from the substrate-binding site to prevent a back-flow of electrons.

Keywords
Cytochrome b561; Ascorbate; Monodehydroascorbate radical; Transmembrane electron transfer; Membrane proteinAsA, ascorbate; CG, chromaffin granule; DEPC, diethylpyrocarbonate; EPR, electron paramagnetic resonance; MALDI-TOF, matrix assisted laser desorptio
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Roles of conserved Arg72 and Tyr71 in the ascorbate-specific transmembrane electron transfer catalyzed by Zea mays cytochrome b561
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 115, Issue 5, May 2013, Pages 497–506
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us