Cloning, genetic organization, and characterization of a structural gene encoding bacillopeptidase F from Bacillus subtilis.
Bacillopeptidase F is an extracellular serine protease that is expressed at the beginning of the stationary phase. To study its structure, regulation of expression, and physiological roles, we have cloned and characterized the structural gene (bpf) encoding this protease from Bacillus subtilis. DNA sequence analysis suggests this protease is synthesized as a preproenzyme (Mr = 92,000). Through processing at both the NH2 and COOH termini, it is gradually converted into various forms with molecular mass ranging from 80 to 48 kDa. Shortening the 3' end of bpf demonstrates that at least 290 amino acid residues from the COOH-terminus of bacillopeptidase F are not required for either catalytic activity or secretion. Bacillopeptidase F exhibits sequence similarity with several serine proteases. Its gene is found immediately downstream from the fts operon which was mapped at 135 degrees on the B. subtilis genetic linkage map. Inactivation of the chromosomal copy of bpf shows no effect on cell growth and sporulation. A triple protease-deficient strain (WB300 with the structural genes for bacillopeptidase F and two other major proteases inactivated) was constructed to serve as a better expression host for the production and secretion of foreign proteins.
Wu
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XC
Department of Biological Sciences, University of Calgary, Alberta, Canada.
Nathoo
S
S
Pang
A S
AS
Carne
T
T
Wong
S L
SL
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D007700:Kinetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D012697:Serine Endopeptidases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D012697:Serine Endopeptidases; C038477:bacillopeptidase F