嗜麦芽寡养单胞菌胞外蛋白酶的化学修饰

Chemical Modification of Extracellular Protease of Stenotrophomonas maltophilia

分别采用苯甲基磺酰氟(PMSF)、对-氯汞苯甲酸(PCMB)、N-乙咪唑(N-AI)、氯胺-T(Ch-T)、N-溴化琥珀亚胺(NBS)、2-巯基乙醇(2-ME)6种化学修饰剂处理嗜麦芽寡养单胞菌胞外蛋白酶,研究酶分子中氨基酸侧链基团与酶活性的关系.结果表明Ch-T,NBS和2-ME能显著抑制酶活性,而N-AI,PMSF和PCMB对酶活性的影响不大,说明蛋氨酸残基、色氨酸残基和二硫键是酶活性的必需基团,而酪氨酸残基、丝氨酸残基和巯基与酶活性无直接关系.同时检测了EDTA和金属离子对酶活性的影响.实验结果证实,EDTA,Mg2+,Ca2+,Hg2+和Cu2+能显著影响酶活性,说明该酶为一种金属蛋白酶.

Extracellular protease was one of the important pathogenic factors of Stenotrophomonas maltophilia. In order to investigate the relationship of structure and function of extracellular protease, the effects of protein modification reagents on the S. maltophilia extracellular protease activity have been studied. The chemical modification of extraceUular protease of S. maltophilia was studied with phenylmethanesulfonyl fluoride （PMSF）, p-chloromercuribenzoate （PCMB）, N-acetylimidazole （N-AI）, chloramine-T （Ch-T）, N-bromosuccinimide （NBS）, 2-mercaptoethanol （2-ME） as the modification reagents. The results indicated that the protease was not affected by N-AI, PMSF and PCMB, but Ch-T, NBS and 2-ME could reduce the activity significantly. It implied that Ser, Tyr and sulfhydryl residues were not activity dependent groups, while Met, Trp and disulfide linkage groups played an important role in the activity of extracellular protease. Meanwhile, EDTA, Mg^2＋, Ca^2＋, Hg^2＋ and Cu^2＋ could reduce the activity obviously, which showed that the enzyme was a metalloproteinase.