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@article{1547016,
author = {Sedláček, Vojtěch and Kučera, Igor},
article_location = {HOBOKEN},
article_number = {1},
doi = {http://dx.doi.org/10.1111/mmi.14260},
keywords = {KINETIC MECHANISM; REDUCED FLAVIN; OXIDATIVE STRESS; SUPEROXIDE ANION; NADH OXIDASE},
language = {eng},
issn = {0950-382X},
journal = {Molecular Microbiology},
title = {Functional and mechanistic characterization of an atypical flavin reductase encoded by the pden_5119 gene in Paracoccus denitrificans},
url = {http://dx.doi.org/10.1111/mmi.14260},
volume = {112},
year = {2019}
}
TY - JOUR
ID - 1547016
AU - Sedláček, Vojtěch - Kučera, Igor
PY - 2019
TI - Functional and mechanistic characterization of an atypical flavin reductase encoded by the pden_5119 gene in Paracoccus denitrificans
JF - Molecular Microbiology
VL - 112
IS - 1
SP - 166-183
EP - 166-183
PB - WILEY
SN - 0950382X
KW - KINETIC MECHANISM
KW - REDUCED FLAVIN
KW - OXIDATIVE STRESS
KW - SUPEROXIDE ANION
KW - NADH OXIDASE
UR - http://dx.doi.org/10.1111/mmi.14260
L2 - http://dx.doi.org/10.1111/mmi.14260
N2 - Pden_5119, annotated as an NADPH-dependent FMN reductase, shows homology to proteins assisting in utilization of alkanesulfonates in other bacteria. Here, we report that inactivation of the pden_5119 gene increased susceptibility to oxidative stress, decreased growth rate and increased growth yield; growth on lower alkanesulfonates as sulfur sources was not specifically influenced. Pden_5119 transcript rose in response to oxidative stressors, respiratory chain inhibitors and terminal oxidase downregulation. Kinetic analysis of a fusion protein suggested a sequential mechanism in which FMN binds first, followed by NADH. The affinity of flavin toward the protein decreased only slightly upon reduction. The observed strong viscosity dependence of k(cat) demonstrated that reduced FMN formed tends to remain bound to the enzyme where it can be re-oxidized by oxygen or, less efficiently, by various artificial electron acceptors. Stopped flow data were consistent with the enzyme-FMN complex being a functional oxidase that conducts the reduction of oxygen by NADH. Hydrogen peroxide was identified as the main product. As shown by isotope effects, hydride transfer occurs from the pro-S C4 position of the nicotinamide ring and partially limits the overall turnover rate. Collectively, our results point to a role for the Pden_5119 protein in maintaining the cellular redox state.
ER -
SEDLÁČEK, Vojtěch a Igor KUČERA. Functional and mechanistic characterization of an atypical flavin reductase encoded by the pden\_{}5119 gene in Paracoccus denitrificans. \textit{Molecular Microbiology}. HOBOKEN: WILEY, 2019, roč.~112, č.~1, s.~166-183. ISSN~0950-382X. Dostupné z: https://dx.doi.org/10.1111/mmi.14260.
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