Characterization of an extracellular protease inhibitor of Bacillus brevis HPD31 and nucleotide sequence of the corresponding gene.
A novel proteinaceous protease inhibitor was isolated from the culture supernatant of Bacillus brevis HPD31. The protease inhibitor of B. brevis (designated BbrPI) was produced extracellularly in multiple forms having at least three different molecular weights. One of them, BbrPI-a, was purified to near homogeneity and only showed inhibitory activity toward serine proteases, such as trypsin, chymotrypsin, and subtilisin. BbrPI was presumed to form a trypsin-inhibitor complex in a molar ratio of 1:1. The inhibitor was found to be heat resistant at neutral and acidic pHs. The gene coding for BbrPI was cloned into Escherichia coli, and its nucleotide sequence was determined. The sequence suggested that BbrPI is produced with a signal peptide of 24 amino acid residues. The amino acid sequence of the protein deduced from the DNA sequence contained the amino acid sequences of amino termini of the inhibitors, a, b, and c, and their putative precursor determined chemically. The molecular weight of the precursor was about 33,000, and the molecular weights of inhibitors a, b, and c were about 22,000, 23,500, and 24,000, respectively. It is presumed that the secreted precursor protein, which is probably inactive, is cleaved by protease into several active protease inhibitor molecules. BbrPI shows no significant homology to the protease inhibitors described previously and is unique in not having any cysteine residues in its molecule.
Shiga
Y
Y
Department of Food Science and Technology, Faculty of Agriculture, Nagoya University, Japan.
Hasegawa
K
K
Tsuboi
A
A
Yamagata
H
H
Udaka
S
S
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000378:metabolism*; D015183:Restriction Mapping
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D011480:Protease Inhibitors; C075183:protease inhibitor, Bacillus brevis