THE PRIMARY STRUCTURE OF THE BACILLUS-AMY LOLIQUEFACIENS INTRACELLULAR SERINE PROTEINASE .3. AMINO-ACID-SEQUENCES OF PEPTIDES FROM THE GLU,ASP-SPECIFIED PROTEINASE HYDROLYSATE - RECONSTRUCTION OF THE INTRACELLULAR SERINE PROTEINASE PRIMARY STRUCTURE

Glu,Asp-specified protease hydrolysate of intracellular serine protein ase (ISP) was separated by ion-exchange chromatography on a sulphocati onite resin followed by HPLC to yield 30 individual peptides. Their se quences, spanning to 243 amino acid residues, were determined by the m anual Edman procedure. Four overlapping fragments were reconstructed b y comparing their sequences with those of tryptic and chymotryptic pep tides. To arrange these fragment in the proteinase polypeptide chain a nd to reconstruct the enzyme's total sequence, additional peptides wer e isolated from the tryptic hydrolysate and analysed. Primary structur e of ISP, corresponding to 297 amino acid residues, was reconstructed. Its comparison with related serine proteinases revealed the following levels of homology: with Bacillus subtilis intracellular serine prote inase, 88%; with secretory subtilisin BPN' produced by B. amyloliquefa ciens, 46%.