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Purification, properties and primary structure of alanine dehydrogenase involved in taurine metabolism in the anaerobe Bilophila wadsworthia.
- Source :
-
Archives of microbiology [Arch Microbiol] 2000 Sep; Vol. 174 (3), pp. 162-7. - Publication Year :
- 2000
-
Abstract
- Alanine dehydrogenase [L-alanine:NAD+ oxidoreductase (deaminating), EC 1.4.1.4.] catalyses the reversible oxidative deamination of L-alanine to pyruvate and, in the anaerobic bacterium Bilophila wadsworthia RZATAU, it is involved in the degradation of taurine (2-aminoethanesulfonate). The enzyme regenerates the amino-group acceptor pyruvate, which is consumed during the transamination of taurine and liberates ammonia, which is one of the degradation end products. Alanine dehydrogenase seems to be induced during growth with taurine. The enzyme was purified about 24-fold to apparent homogeneity in a three-step purification. SDS-PAGE revealed a single protein band with a molecular mass of 42 kDa. The apparent molecular mass of the native enzyme was 273 kDa, as determined by gel filtration chromatography, suggesting a homo-hexameric structure. The N-terminal amino acid sequence was determined. The pH optimum was pH 9.0 for reductive amination of pyruvate and pH 9.0-11.5 for oxidative deamination of alanine. The apparent Km values for alanine, NAD+, pyruvate, ammonia and NADH were 1.6, 0.15, 1.1, 31 and 0.04 mM, respectively. The alanine dehydrogenase gene was sequenced. The deduced amino acid sequence corresponded to a size of 39.9 kDa and was very similar to that of the alanine dehydrogenase from Bacillus subtilis.
- Subjects :
- Alanine Dehydrogenase
Amino Acid Sequence
Bacteria, Anaerobic genetics
Bacteria, Anaerobic growth & development
Catalysis
DNA, Bacterial genetics
Deltaproteobacteria genetics
Deltaproteobacteria growth & development
Kinetics
Molecular Sequence Data
Sequence Analysis, DNA
Amino Acid Oxidoreductases chemistry
Amino Acid Oxidoreductases genetics
Amino Acid Oxidoreductases isolation & purification
Amino Acid Oxidoreductases metabolism
Bacteria, Anaerobic enzymology
Deltaproteobacteria enzymology
Taurine metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0302-8933
- Volume :
- 174
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- Archives of microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 11041346
- Full Text :
- https://doi.org/10.1007/s002030000190