金黄色葡萄球菌核酸酶A在温控双表达载体中的表达及其活性分析

Expression and activity analysis of staphylococcal nuclease A in a thermo-inducible dual expressing vector

菌蜕制备时,金黄色葡萄球菌核酸酶A(Staphylococcal nuclease A,SNA)在温控单表达系统中表达时,对宿主菌的灭活效率比其在双表达系统中与基因E共表达时的灭活效率高近一个数量级。为探究导致这种差异的可能机制,将SNA克隆至一温控双表达载体进行诱导表达并对其活性进行综合分析。结果显示,胞内和胞外产物均具有核酸酶活性,分别在升温诱导30 min和90 min后被检出,对宿主菌的4 h灭活率为99.7%,宿主基因组亦被成功降解。这表明,SNA在温控双表达载体中的表达仍能如在单表达载体中一样有效灭活细菌,裂解基因E和SNA的共表达可能才是造成上述差异的主要原因。

The efficiency of inactivating bacterial hosts of staphylococcal nuclease A（SNA） is at one order of magnitude higher when it is expressed in a thermo-inducible single expression system than in a dual expressing system combined with simultaneously expressing lysis gene E for bacterial ghost preparation. To explore possible mechanisms causing this difference,SNA was cloned into a thermo-inducible dual expressing vector and its enzymatic activity was analyzed after induction. Results showed that intracellular and extracellular products of SNA were detectable at 30 min and 90 min,respectively after induction and all possess nuclease activity. The percentage of inactivated host bacteria was 99. 7% 4 h after induction,with the host genomic DNA successfully degraded. These indicated that the expression of SNA in a thermo-inducible dual expressing vector without simultaneously expression of lysis gene E could still kill the host strain as efficiently as in a single expressing system. The co-expression of SNA and lysis gene E in the dual expressing vector might be the major reason resulting in lower inactivation efficiency of SNA.