Thiolsubtilisin as an instrument for peptide synthesis. Preparation and properties.
A convenient procedure for thiolsubtilisin purification from an admixture of subtilisin involving affinity chromatography on bacitracin-Sepharose is presented. Thiolsubtilisin activity was measured by hydrolysis of p-nitrophenyl acetate, p-nitroanilide-peptide (Glp-Ala-Ala-Leu-pNA), and azocasein. The thiolenzyme catalyzes peptide synthesis. Under these conditions only activated peptide esters, e.g., p-chlorophenyl, N-hydroxysuccinimide, or p-nitrophenyl esters form peptide bonds during interaction with appropriate nucleophiles such as peptides and their derivatives and amino acid amides.
Kolobanova
S V
SV
School of Chemistry, Lomonosov Moscow State University, Russia.
Lysogorskaya
E N
EN
Filippova IYu
Anisimova
V V
VV
Oksenoit
E S
ES
Stepanov
V M
VM
D000596:Amino Acids / Q000032:analysis; D001412:Bacillus subtilis / Q000201:enzymology*; D001414:Bacitracin; D002364:Caseins / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel; D006868:Hydrolysis; D009596:Nitrophenols; D010452:Peptide Biosynthesis; D010455:Peptides / Q000737:chemistry; D012685:Sepharose; D013379:Substrate Specificity; D013381:Subtilisins / Q000302:isolation & purification* / Q000378:metabolism*
D000596:Amino Acids; D002364:Caseins; D009596:Nitrophenols; D010455:Peptides; C026018:azocasein; C042614:bacitracin-Sepharose complex; D001414:Bacitracin; C008642:4-nitrophenyl acetate; D012685:Sepharose; D013381:Subtilisins; C022743:thiolsubtilisins