Characterization of endogenous pyridoxal 5'-phosphate-dependent alanine racemase from Bacillus pseudofirmus OF4.
J Biosci Bioeng
; 107(3): 225-9, 2009 Mar.
Article
en En
| MEDLINE
| ID: mdl-19269582
ABSTRACT
An open reading frame of 1100 bp in the partially sequenced genome sequence of alkaliphilic Bacillus pseudofirmus OF4 was identified as a putative alanine racemase gene (dadX(OF4)), which was cloned and expressed in Escherichia coli BL21 (DE3). The encoded protein DadX(OF4) was purified to homogeneity by His(6)-tag affinity column, gel filtration and ion-exchange chromatography. The amino acid sequence has highest identity with the known alanine racemase from Oceanobacillus iheyensis HTE831 (48%). The protein was a dimeric, endogenous PLP-dependent enzyme, which was demonstrated by absorption spectra and enzyme activity with or without PLP. The racemization temperature optimum was 40 degrees C and the optimal pH was 10.5. The kinetic parameters K(m) and V(max) at 40 degrees C of alanine racemase, determined by HPLC analysis, were 41.79 mM, 10,500 units/mg for L-alanine and 14.91 mM, 3708 units/mg for D-alanine, respectively.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Fosfato de Piridoxal
/
Bacillus
/
Proteínas Bacterianas
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Alanina Racemasa
Idioma:
En
Revista:
J Biosci Bioeng
Asunto de la revista:
ENGENHARIA BIOMEDICA
/
MICROBIOLOGIA
Año:
2009
Tipo del documento:
Article
País de afiliación:
China