Molecular cloning and nucleotide sequence of the 90k serine protease gene, hspK, from Bacillus subtilis (natto) No. 16.
We previously reported purification and characterization of a 90k serine protease with pI 3.9 from Bacillus subtilis (natto) No. 16 [Kato et al. 1992 Biosci Biotechnol Biochem 56:1166]. The enzyme showed different and unique substrate specificity towards the oxidized B-chain of insulin from those of well-known bacterial serine proteases from Bacillus subtilisins. The structural gene, hspK, for the 90k serine protease was cloned and sequenced. The cloned DNA fragment contained a single open reading frame of 4302 bp coding a protein of 1433 amino acid residues. The deduced amino acid sequence of the 90k-protease indicated the presence of a typical signal sequence of the first 30 amino acids region and that there was a pro-sequence of 164 amino acid residues after the signal sequence. The mature region of the 90k-protease started from position 195 of amino acid residue, and the following peptide consisted of 1239 amino acid residues with a molecular weight of 133k. It might be a precursor protein of the 90k-protease, and the C-terminal region of 43k might be degraded to a mature protein from the precursor protein. The catalytic triad was thought to consist of Asp33, His81, and Ser259 from comparison of the amino acid sequence of the 90k-protease with those of the other bacterial serine proteases. The high-molecular-weight serine protease, the 90k-protease, may be an ancient form of bacterial serine proteases.
Department of Applied Biological Chemistry, Faculty of Agriculture, Tohoku University, Sendai, Japan.
Y
Y
Yamagata
R
R
Abe
Y
Y
Fujita
E
E
Ichishima
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D005798:Genes, Bacterial / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics; D015183:Restriction Mapping; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013381:Subtilisins / Q000737:chemistry / Q000235:genetics*
D011498:Protein Precursors; D013381:Subtilisins