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Title: Protection of the Queuosine Biosynthesis Enzyme QueF from Irreversible Oxidation by a Conserved Intramolecular Disulfide

Journal Article · · Biomolecules
DOI:https://doi.org/10.3390/biom7010030· OSTI ID:1628313
 [1];  [2];  [2];  [2];  [1];  [2];  [3];  [1]
  1. Western University of Health Sciences, Pomona, CA (United States)
  2. Portland State Univ., OR (United States)
  3. San Diego State Univ., CA (United States)
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QueF enzymes catalyze the nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reduction of the nitrile group of 7-cyano-7-deazaguanine (preQ0) to 7-aminomethyl-7-deazaguanine (preQ1) in the biosynthetic pathway to the tRNA modified nucleoside queuosine. The QueF-catalyzed reaction includes formation of a covalent thioimide intermediate with a conserved active site cysteine that is prone to oxidation in vivo. Here, we report the crystal structure of a mutant of Bacillus subtilis QueF, which reveals an unanticipated intramolecular disulfide formed between the catalytic Cys55 and a conserved Cys99 located near the active site. This structure is more symmetric than the substrate-bound structure and exhibits major rearrangement of the loops responsible for substrate binding. Mutation of Cys99 to Ala/Ser does not compromise enzyme activity, indicating that the disulfide does not play a catalytic role. Peroxide-induced inactivation of the wild-type enzyme is reversible with thioredoxin, while such inactivation of the Cys99Ala/Ser mutants is irreversible, consistent with protection of Cys55 from irreversible oxidation by disulfide formation with Cys99. Conservation of the cysteine pair, and the reported in vivo interaction of QueF with the thioredoxin-like hydroperoxide reductase AhpC in Escherichia coli suggest that regulation by the thioredoxin disulfide-thiol exchange system may constitute a general mechanism for protection of QueF from oxidative stress in vivo.

Research Organization:
SLAC National Accelerator Laboratory (SLAC), Menlo Park, CA (United States). Stanford Synchrotron Radiation Lightsource (SSRL)
Sponsoring Organization:
USDOE Office of Science (SC), Basic Energy Sciences (BES); USDOE Office of Science (SC), Biological and Environmental Research (BER); National Science Foundation (NSF); National Institutes of Health (NIH); National Institute of General Medical Sciences (NIGMS)
Grant/Contract Number:
AC02-76SF00515; CHE-1309323; GM110588; GM70641; P41GM103393
OSTI ID:
1628313
Journal Information:
Biomolecules, Vol. 7, Issue 4; ISSN 2218-273X
Publisher:
MDPICopyright Statement
Country of Publication:
United States
Language:
English
Citation Metrics:
Cited by: 8 works
Citation information provided by
Web of Science

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Cited By (4)

Allosteric disulphide bonds as reversible mechano-sensitive switches that control protein functions in the vasculature journal May 2019
Interplay of nucleophilic catalysis with proton transfer in the nitrile reductase QueF from Escherichia coli journal January 2019
Evidence of a sequestered imine intermediate during reduction of nitrile to amine by the nitrile reductase QueF from Escherichia coli journal January 2018
Allosteric disulfides: Sophisticated molecular structures enabling flexible protein regulation journal January 2019

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59 BASIC BIOLOGICAL SCIENCES
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