灰色链霉菌RX-17溶菌酶R1的纯化及性质研究

通过硫酸铵分级沉淀，CM-Sephadex C50、CM-Sepharose Fast Flow离子交换层析及Sephadex G-75凝胶过滤层析，从灰色链霉菌（Streptomyces griseus）RX17的发酵上清液中得到了电泳纯的溶菌酶R1，回收率6.89%。测得该酶分子量和等电点分别为16.8kD和9.10，作用于变链球菌（Streptococcus mutans）Ingbritt的最适温度和pH分别为70℃和6.6。R1酶在50℃以下及pH6～9的范围内保持稳定，60℃保温1h，残存酶活20.3％。Mg2+对酶有激活作用，而Zn2+、Cu2+、Fe2+、Cd2+、Pb2+则使酶完全丧失活性，螯合剂、盐酸羟胺、碘乙酸抑制酶活，β-巯基乙醇及表面活性剂则对溶菌有部分促进作用。R1酶溶菌谱广泛，对多种卵清溶菌酶不能作用的G+、G细菌均有溶解能力，对变链球菌、金黄色葡萄球菌（Staphylococcus aureus）、乳杆菌(Lactobacillus)等则呈现高活性。

Purification and Some Properties of Bacteriolytic Enzyme R1 from Streptomyces griseus RX-17

Bacteriolytic enzyme R1 was purified to electrophoretic homogeneity with the recovery of 6.89% activity by ammonium sulfate precipitation, CM-Sephadex C - 50, CM-Sepharose Fast Flow and Sephadex G-75 chromatography from the culture supernatant of Streptomyces griseus RX-17. The molecular weight and PI of R1 were 16.8 kD and 9.10. The optimal temperature and pH for R1 against Streptococcus mutans Ingbritt were 70 degrees C and 6.6, respectively. Below 50 degrees C and at range pH 6 - 10, R1 was stable. While treated at 60 degrees C for 1 hour, the residual activity was only about 20.3%. Zn2+, Cu2+, Fe2+, Cd2+ and Pb2+ could completely inactivate the enzyme. Chelating agents, hydroxylamine hydrochloriae, Monoiodoacetic acid inhibited the lytic activity against Streptococcus mutans Ingbritt, whereas Mg2+, 2-Mercaptoethanol and some surfactants could stimulate the activity. The enzyme had a broad bacteriolytic spectrum against many G+, G- bacteria which were resistant to egg-white lysozyme. Especially high activity was shown on Streptococcus mutans, Staphylococcus aureus and Lactobaillus.