Isolation and analysis of metA, a methionine biosynthetic gene encoding homoserine acetyltransferase in corynebacterium glutamicum.
The metA gene encoding homoserine acetyltransferase, the first enzyme of the methionine biosynthetic pathway, was isolated from a pMT1-based corynebacterium glutamicum gene library via complementation of an Escherichia coli metA mutant. A DNA-sequence analysis of the cloned DNA is identified an open-reading frame of 1,137 bp which encodes a protein with the molecular weight of 41,380 comprising 379 amino acids. The putative protein product showed good amino acid-sequence homology to its counterpart in other organisms. The internal fragment of the cloned DNA was successfully used to disrupt chromosomal metA, demonstrating the identity of the cloned gene. The C. glutamicum metA mutant lost the ability to grow on glucose minimal medium supplemented with homoserine. However, the mutant could grow on a minimal medium supplemented with cystathionine, demonstrating that C. glutamicum uses the cystathionine route to synthesize methionine. Introduction of a plasmid carrying cloned metA into C. glutamicum resulted in a 10-fold increase in enzyme activities and expression of a protein product of M(r) 41,000, which agrees with the sequence data and is similar in size to those of other homoserine acetyltransferases. Unlike E. coli whose metA product uses succinyl coenzyme A as a substrate, the cloned metA gene produced homoserine acetyltransferase which uses only acetyl coenzyme A as the acyl donor.
Park
S D
SD
Graduate School of Biotechnology, Korea University, Seoul, Korea.
Lee
J Y
JY
Kim
Y
Y
Kim
J H
JH
Lee
H S
HS
D000123:Acetyltransferases / Q000235:genetics*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002876:Chromosomes, Bacterial / Q000235:genetics; D003001:Cloning, Molecular; D003352:Corynebacterium / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification*; D015870:Gene Expression / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D015971:Gene Expression Regulation, Enzymologic; D005798:Genes, Bacterial / Q000235:genetics*; D008715:Methionine / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics; D009154:Mutation / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008715:Methionine; D000123:Acetyltransferases; C021369:homoserine O-acetyltransferase