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B6db references: 1110001112

type Journal Article
authors Chen, C-C. Chou, T-L. Lee, C-Y.
title Cloning, expression and characterization of L-aspartate beta-decarboxylase gene from Alcaligenes faecalis CCRC 11585
journal J Industrial Microbiol Biotechnol
Activity 4.1.1.12
Family 4.1.1.12
sel selected
ui 1110001112
year (2000)
volume 25
pages 132-140
 
abstract L-Aspartate beta-decarboxylase (Asd) is an important enzyme to produce L-alanine and D-aspartate. The genomic library of Alcaligenes faecalis CCRC 11585 was cloned into pBK-CMV and transformed into Escherichia coli. One clone, which carried the asd gene and expressed Asd activity, was isolated and chosen for further study. PBK-asdAE1 was subcloned and its sequence anal. revealed an open reading frame, consisting of 1599 bp, that encodes a 533-amino-acid polypeptide. The nucleotide sequence of the asd gene from A. faecalis CCRC 11585 (asdA) showed 84% identity with that from Pseudomonas dacunhae CCRC 12623, and the amino acid sequence showed 93% identity. The amino acid sequence of the AsdA showed 51-58% homol. with various aminotransferases. Alignment of the AsdA with several aspartate or tyrosine aminotransferases revealed 17 conserved amino acids that appeared in most of the conserved amino acid residues within the pyridoxal-5'-phosphate (PLP) binding domains of aminotransferases. Furthermore, the asdA gene was cloned into expression vector pET-21a and transformed into E. coli BL21 (DE3). A protein band sized at 61 kDa is present on the SDS-PAGE gel from the intracellular sol. from of E. coli BL21 (DE3)/pET-asdA. The specific activities of the pET-AsdA purified by using His-Bind chromatog. is 215 U/mg at 45°C and pH 5.0, which is 1000-fold higher than that of the A. faecalis crude ext. This is the first report of an asdA gene sequence from A. faecalis and represents the potential application of a recombinant AsdA for prodn. of L-alanine or D-aspartic acid.
last changed 2007/12/21 17:44

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