Subtilisin BPN proteinase

The sequence of aqualysin Il6) (AQU) is shown compared with those of proteinase Kl7) (PRO), thermitase22 (THE), subtilisin BPN 18 (BPN), and subtilisin El9) (E). Identical amino acids with those of aqualysin I are shown by hyphen (—). Open space is the position where a corresponding amino acid is absent. The numbering above the sequences refers to aqualysin I, and that below the sequences to subtilisins. Asterisks indicate the active-site residues, Asp, His, and Ser. 

Three-dimensional structures of four subtilisin-type enzymes, subtilisin BPN, 36373 subtilisin Carlsberg,37,383 thermitase,39,403 and proteinase K,40,413 are known, but that of aqualysin I has not yet been determined. The Ca atoms of the known structures were superimposed to obtain maximal overlap of the backbone structures, and large parts of all four structures overlap very well (Fig. 12.3) 423 On the basis of such analyses, structurally equivalent core residues (194 residues) are identified, and higher sequence identity was found to correspond to a closer overlap of mainchain atoms in the core (Table 12.1).423... 

Recent data of Fraser, Corstorphine, and Zammit using trypsin and proteinase K treatment of intact mitochondria and outer-membrane-ruptured mitochondria suggest that both the active site and the malonyl-CoA binding site are exposed on the cytosolic side of the membrane and that CPT-I has two transmembrane domains. Thus, we have conducted additional studies of the topology of CPT-I using intact hepatic mitochondria and isolated mitochondrial outer membranes with Nagarse (subtilisin BPN ), papain, and trypsin using a variety of incubation conditions. 

Pepsin, pepsin-like enzymes, chymosin, rennin, and other acid proteinases have an activity optimum at pH 2.0-3.5 papain, trypsin, chymotrypsin, and similar enzymes are most active at neutral pH (pH 6-8). Subtilisin BPN, pancreatic elastase, leucine... 

Uehara, Y., Tonomura, B., and Hiromi, K. (1978). Direct fluoro-metric determination of a dissociation constant as low as 10 0 M for the subtilisin BPN —protein proteinase inhibitor (StrepTotnyces subtilisin inhibitor) complex by a single photon counting technique. J. Biochem. (Tokyo) 84, 1195-1202. 

Fig. 3. Comparison of sequences of thermitase, subtilisins and proteinase K. Top numbering - thermitase, bottom numbering - subtilisin BPN . 

Subtilisin (EC 3.4.21.4) an extracellular, single chain, alkaline serine protease from Bacillus subtilis and related species. S. are known from four different species of Bacillus S. Carlsberg (274 amino acid residues, M, 27,277), S. BPN (275 amino acid residues, M, 27,537), S. Novo (identical with S.BPN ) and S. amylosacchariticus (275 amino acid residues, M, 27671). The observed sequence differences between different S. represent conservative substitutions and are limited to the surface amino acids. Like the pancreatic proteinases, S. has catalytic Ser22i, His64 and Asnjj residues, but it is structurally very different from the other serine proteases, e. g. the active center of S. is -Thr-Ser-Met-, whereas that of the pancreatic enzymes is -Asp-Ser-Gly- pancreatic enzymes contain 4- disulfide bridges, whereas S. contains none S. contains 31 % a-helical structure and 3 spatially separated domains, whereas the pancreatic enzymes have 10-20% a-helical structure and a high content of p-structures in both types, the active center is a substrate cleft. S. also have a broader substrate specificity than the pancreatic enzymes. This is a notable example of the convergent evolution of catalytic activity in two structurally completely different classes of proteins. S. is used in the structural elucidation... 

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