Pyruvate:Quinone Oxidoreductase from Corynebacterium glutamicum: Purification and Biochemical Characterization
AUTOR(ES)
Schreiner, Mark E.
FONTE
American Society for Microbiology
RESUMO
Pyruvate:quinone oxidoreductase catalyzes the oxidative decarboxylation of pyruvate to acetate and CO2 with a quinone as the physiological electron acceptor. So far, this enzyme activity has been found only in Escherichia coli. Using 2,6-dichloroindophenol as an artificial electron acceptor, we detected pyruvate:quinone oxidoreductase activity in cell extracts of the amino acid producer Corynebacterium glutamicum. The activity was highest (0.055 ± 0.005 U/mg of protein) in cells grown on complex medium and about threefold lower when the cells were grown on medium containing glucose, pyruvate, or acetate as the carbon source. From wild-type C. glutamicum, the pyruvate:quinone oxidoreductase was purified about 180-fold to homogeneity in four steps and subjected to biochemical analysis. The enzyme is a flavoprotein, has a molecular mass of about 232 kDa, and consists of four identical subunits of about 62 kDa. It was activated by Triton X-100, phosphatidylglycerol, and dipalmitoyl-phosphatidylglycerol, and the substrates were pyruvate (kcat = 37.8 ± 3 s−1; Km = 30 ± 3 mM) and 2-oxobutyrate (kcat = 33.2 ± 3 s−1; Km = 90 ± 8 mM). Thiamine pyrophosphate (Km = 1 μM) and certain divalent metal ions such as Mg2+ (Km = 29 μM), Mn2+ (Km = 2 μM), and Co2+ (Km = 11 μM) served as cofactors. In addition to several dyes (2,6-dichloroindophenol, p-iodonitrotetrazolium violet, and nitroblue tetrazolium), menadione (Km = 106 μM) was efficiently reduced by the purified pyruvate:quinone oxidoreductase, indicating that a naphthoquinone may be the physiological electron acceptor of this enzyme in C. glutamicum.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=545707Documentos Relacionados
- Regulation of Quinone Oxidoreductase by the Redox-sensing Transcriptional Regulator QorR in Corynebacterium glutamicum*
- Structural and functional analysis of pyruvate kinase from Corynebacterium glutamicum.
- Purification and biochemical characterization of pyruvate oxidase from Lactobacillus plantarum.
- Characterization of the isocitrate lyase gene from Corynebacterium glutamicum and biochemical analysis of the enzyme.
- Export of l-Isoleucine from Corynebacterium glutamicum: a Two-Gene-Encoded Member of a New Translocator Family