Kim N, Kim S I
Food Chemistry and Physics Division, Korea Food Research Institute, Bundang-ku, Songnam-si, Kyonggi-do.
Can J Microbiol. 1994 Feb;40(2):120-6. doi: 10.1139/m94-019.
An extracellular endopeptidase (proteinase) from Serratia marcescens (Serratia marcescens extracellular proteinase, EC 3.4.24.4), purified to homogeneity, was analyzed for enzyme properties. The enzyme has a polypeptide chain molecular mass of 52 kDa as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme has an optimal temperature of 40 degrees C and an optimal pH of 7.0. Enzyme activity was enhanced over two times by the addition of Ca2+ and Mg2+ ions and eliminated almost completely by the presence of 0.2% SDS. The enzyme has broad substrate specificity and contains neither cysteine nor methionine. Low homology was found between the NH2-terminal amino acid sequence of the enzyme of this study and the NH2-terminal sequence of a proteinase from another strain of S. marcescens. Chemical modification with N-bromosuccinimide, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, and 8-anilino-1-naphthalene sulfonic acid and by photooxidation with methylene blue reduced enzyme activity considerably. The enzyme was shown to have broad peptide bond specificity judging from the contribution of 11 amino acids to the carboxyl side of the peptide bonds hydrolyzed.
对一株粘质沙雷氏菌来源的细胞外内肽酶（蛋白酶）（粘质沙雷氏菌细胞外蛋白酶，EC 3.4.24.4）进行了纯化直至达到同质，并对其酶学性质进行了分析。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）测定，该酶的多肽链分子量为52 kDa。该酶的最适温度为40℃，最适pH为7.0。添加Ca2+和Mg2+离子可使酶活性提高两倍以上，而0.2% SDS的存在几乎可使酶活性完全消除。该酶具有广泛的底物特异性，且不含半胱氨酸和甲硫氨酸。本研究中该酶的NH2末端氨基酸序列与另一株粘质沙雷氏菌蛋白酶的NH2末端序列之间的同源性较低。用N-溴代琥珀酰亚胺、1-乙基-3-(3-二甲基氨基丙基)-碳二亚胺和8-苯胺基-1-萘磺酸进行化学修饰，以及用亚甲蓝进行光氧化，均会使酶活性大幅降低。从水解的肽键羧基侧11种氨基酸的贡献来看，该酶具有广泛的肽键特异性。
