Cloning and sequence analysis of the pfl gene encoding pyruvate formate-lyase from Streptococcus mutans.
We have isolated a sorbitol-negative mutant of Streptococcus mutans GS-5 following random mutagenesis with plasmid pVA891 clone banks. This mutant did not metabolize sorbitol anaerobically but did so aerobically. A 10-kb chromosomal DNA fragment flanking the pVA891 insertion was deleted in this mutant. The corresponding region from the parental strain GS-5 was then recovered by a marker rescue method with Escherichia coli. The pyruvate formate-lyase gene, pfl, was identified within a 3-kb PstI-XbaI fragment located in the middle of the deleted region of the chromosome, and its inactivation in S. mutans produced the same sorbitol-negative phenotype. Nucleotide sequence analysis of the pfl gene revealed a 2.3-kb open reading frame (ORF) preceded by potential ribosome-binding and promoter-like sequences. The ORF specified a putative protein of 775 amino acid residues with a calculated molecular weight of 87,533. The amino acid sequence deduced from the ORF exhibited significant similarity to that of the E. coli pfl gene.
Department of Biochemistry, Tokyo Dental College, Chiba City, Japan.
Y
Y
Yamamoto
Y
Y
Sato
S
S
Takahashi-Abbe
K
K
Abbe
T
T
Yamada
H
H
Kizaki
D000123:Acetyltransferases / Q000737:chemistry / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D016366:Open Reading Frames; D013295:Streptococcus mutans / Q000201:enzymology* / Q000235:genetics
D000123:Acetyltransferases; C022560:formate C-acetyltransferase