The structural characterization of subtilisin mesoscale clusters, which were previously shown to induce supramolecular order in biocatalytic self-assembly of Fmoc-dipeptides, was carried out by synchrotron small-angle X-ray, dynamic, and static light scattering measurements. Subtilisin molecules self-assemble to form supramolecular structures in phosphate buffer solutions. Structural arrangemen...

To ascertain whether position 131 of a mesophilic protease, subtilisin BPN', is a potential critical site for cold adaptation as screened by evolutionary engineering (S. Taguchi, A. Ozaki, and H. Momose, Appl. Environ. Microbiol. 64:492-495, 1998), a full set of subtilisin BPN' mutants with mutations at position 131 was constructed by site-saturation mutagenesis. All mutated enzymes were measur...

AIMS The nature of exopolymers involved in the adhesion of a marine biofilm-forming bacterium Pseudoalteromonas sp. D41 was investigated to evaluate and understand the antifouling potential of subtilisin. METHODS AND RESULTS The exopolymers of D41 produced by fermentation were analysed by FTIR and SDS-PAGE showing the presence of polysaccharides, glycoproteins and proteins. A high content of ...

Subtilisin DY is very resistant to the denaturing action of urea: the conformational properties are not affected up to 4.5 M-urea, and even in the presence of 8 M-urea there is only a slow loss of ordered structure and caseinolytic activity. C.d. and fluorescence-emission studies also show that this proteinase is stable in the 5.5-10.0 pH range, whereas below pH 5.5 a sharp denaturation occurs ...

Sequential rounds of error-prone PCR to introduce random mutations and screening of the resultant mutant libraries have been used to enhance the total catalytic activity of subtilisin E significantly in a non-natural environment, aqueous dimethylformamide (DMF). Seven DNA substitutions coding for three new amino acid substitutions were identified in a mutant isolated after two additional genera...

Random mutagenesis has been used to engineer the protease subtilisin E to function in a highly nonnatural environment--high concentrations of a polar organic solvent. Sequential rounds of mutagenesis and screening have yielded a variant (PC3) that hydrolyzes a peptide substrate 256 times more efficiently than wild-type subtilisin in 60% dimethylformamide. PC3 subtilisin E and other variants con...

A three-dimensional structural model of nattokinase (NK) from Bacillus natto was constructed by homology modeling. High-resolution X-ray structures of Subtilisin BPN' (SB), Subtilisin Carlsberg (SC), Subtilisin E (SE) and Subtilisin Savinase (SS), four proteins with sequential, structural and functional homology were used as templates. Initial models of NK were built by MODELLER and analyzed by...

All preparations of ribonuclease-S so far reported have employed limited digestion of ribonuclease-A with subtilisin,' a bacterial proteinase from B. subtilis described by Giintelberg and Ottesen.' The original supply of this latter enzyme is now exhausted. Another sample of a crystalline proteinase from B. subtilis (NOVO Enzyme) has been shown to differ from the original in several respects,' ...