Aldehyde oxidoreductase activity Desulfovibrio alaskensis NCIMB 13491. EPR assignment of the proximal [2Fe-2S] cluster to the Mo site

Susana L. A. Andrade, Carlos D. Brondino, Maria J. Feio, Isabel Moura, José J. G. Moura

Research output: Contribution to journalArticlepeer-review

31 Citations (Scopus)

Abstract

A novel molybdenum iron-sulfur-containing aldehyde oxidoreductase (AOR) belonging to the xanthine oxidase family was isolated and characterized from the sulfate-reducing bacterium Desulfovibrio alaskensis NCIMB 13491, a strain isolated from a soured oil reservoir in Purdu Bay, Alaska. D. alaskensis AOR is closely related to other AORs isolated from the Desulfovibrio genus. The protein is a 97-kDa homodimer, with 0.6 ± 0.1 Mo, 3.6 ± 0.1 Fe and 0.9 ± 0.1 pterin cytosine dinucleotides per monomer. The enzyme catalyses the oxidation of aldehydes to their carboxylic acid form, following simple Michaelis-Menten kinetics, with the following parameters (for benzaldehyde): K(m)/(app) = 6.65 μM: (app) = 13.12 μM·min-1; k(cat)/(app) = 0.96 s-1. Three different EPR signals were recorded upon long reduction of the protein with excess dithionite: an almost axial signal split by hyperfine interaction with one proton associated with Mo(V) species and two rhombic signals with EPR parameters and relaxation behavior typical of [2Fe-2S] clusters termed Fe/S I and Fe/S II, respectively. EPR results reveal the existence of magnetic interactions between Mo(V) and one of the Fe/S clusters, as well as between the two Fe/S clusters. Redox titration monitored by EPR yielded midpoint redox potentials of -275 and -325 mV for the Fe/S I and Fe/S II, respectively. The redox potential gap between the two clusters is large enough to obtain differentiated populations of these paramagnetic centers. This fact, together with the observed interactions among paramagnetic centers, was used to assign the EPR-distinguishable Fe/S I and Fe/S II to those seen in the reported crystal structures of homologous enzymes.

Original languageEnglish
Pages (from-to)2054-2061
Number of pages8
JournalEuropean Journal Of Biochemistry
Volume267
Issue number7
DOIs
Publication statusPublished - 14 Apr 2000

Keywords

  • Aldehyde oxidoreductase
  • Desulfovibrio alaskensis
  • Molybdenum hydroxylases
  • Xanthine oxidase family

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