RESUMEN
The milk-clotting protease from Bacillus mesentericus strain 76 is free of carbohydrate and lacks cysteine and cystine. The amino acid composition indicates a single peptide chain with 304 residues. Five amino acids--aspartic acid, threonine, serine, glycine and alanine--represent one-half of the total residues. The enzyme contains 35 aromatic amino acids and 103 ionic amino acids. The observed constant value of the ratio Menzyme:Azinc is 1:1 for active and acid denatured enzyme preparations, indicating that 0.1 M acetic acid causes denaturation of the enzyme but it can not eliminate the zinc ions.
Asunto(s)
Ácido Aspártico Endopeptidasas , Bacillus/enzimología , Endopeptidasas/aislamiento & purificación , Aminoácidos/análisis , Carbohidratos/análisis , Espectrofotometría Ultravioleta , Zinc/análisisRESUMEN
A two-step procedure for the isolation of pure enzyme from Bacillus mesentericus strain 76 is developed. Ion-exchange chromatography on CM-Sephadex-C50 is used as a second step after precipitation from ethanol. The pure enzyme preparation, obtained after gel filtration on Sephadex-G25 of the ion-exchange chromatographically separated product, possess the highest specific activity achieved. The homogeneity of the final preparation is proved by determination of the N-terminal amino acid (Arg) and by SDS-PAGDE (single disc).
