金黄色葡萄球菌ltaS突变株的构建与eLtaS蛋白的表达

摘要

目的 通过构建金黄色葡萄球菌(SAU)ltaS突变株以及体外重组表达其胞外形式的eLtaS蛋白,研究LtaS以及eLtaS蛋白在SAU中的生物学功能.方法 利用同源重组的方法构建SAU 8325-4来源的ltaS缺失突变菌株,并对其生长以及外毒素水平进行检测;同时,在大肠杆菌中,体外重组表达并纯化eLtaS蛋白,继而观察其对SAU生长的影响.结果在SAU 8325-4中成功构建了ltaS缺失突变菌株,并发现ltaS突变株生长较野生型菌株缓慢;通过表达纯化成功获得高纯度的eLtaS蛋白,其与突变株共培养后对突变株的生长缓慢现象没有回复作用.结论 LtaS蛋白在SAU侵袭机体的过程中发挥着重要的功能,其胞外形式的eLtaS蛋白可能未参与脂磷壁酸(lipoteichoic acid,LTA)的合成.%Objective To investigate the biological function of LtaS and its extracellular form eLtaS in Staphylococcus aureus ( S. Aureus )by constructing the ItaS mutant from S. Aureus 8325-4, expressing and purifying the eLtaS protein. Methods ItaS Depletion mutant was constructed by homologous recombination in S. Aureus 8325-4. The growth curve and the level of its extra-toxins production were measured. The recombinant eLtaS was expressed and purified in vitro by pET-28a vector, and the effect of eLtaS on the growth of S. Aureus was detected. Results The ItaS mutant was successfully constructed in S. Aureus 8325-4. Compared with the wild-type strain, the growth rate of the mutant was inhibited and the level of extra-toxins of the ItaS mutant was decreased. We found that eLtaS could not restore the growth of the ItaS mutant. Conclusion LtaS plays an important role in the growth and pathogenicity of S. Aureus, while the extracellular protein eLtaS may not be involved in the synthesis of lipoteichoic acid.