嗜碱菌Alkalimonas amylolytica N10在不同pH条件下的膜蛋白差异表达研究

Study on the membrane proteins of Alkalimonas amylolytica N10 expressed at different pH environment

为了探讨嗜碱菌的嗜碱机制，对一株新型专性嗜碱菌（Alkalimonas amylolytico N10）在不同pH条件下的差异膜蛋白质组进行了初步的研究。在3种pH条件下（pH值分别为8．4、9．4和10．4）培养的该菌的膜蛋白，通过8％-20％的梯度SDS—PAGE进行分离。经胶图图像和统计计算，7条电泳条带的相对染色强度随培养液的pH值变化而改变，其中仅有一条条带强度随pH值增高而增加。这些条带经胰蛋白酶胶内消化和高效液相色谱-电喷雾离子阱串联质谱（LC—Ms／Ms）分析，共鉴定出了12种蛋白质。其中4种膜蛋白已有报道直接或间接参与细胞pH稳态的保持，其他几种蛋白则是首次发现其表达水平与生活环境的pH变化可能相关。

The research on the membrane proteins expressed in alkaliphiles at different pH media will be benefit to understand the molecular mechanisms how they survive from such harsh alkaline environment. A preliminary investigation to the differential membrane proteomes from Alkalimonas amylolytica N10 （N10）, a novel alkaliphilic bacterium, is reported herein. Upon the growth curves of N10 at different pH environment, three pH media, 8.4, 9.4 and I0.4, respectively, were employed to culture N10 followed by membrane preparation using ultracentrifuge and membrane proteins separation by 8% - 20% gradient SDS-PAGE. With carefully image analysis and statistical calculation, 7 stained bands were confirmed the alterations of relative optimal intensities corresponding with pH environment. Furthermore these bands were digested in-gel by trypsin and delivered to LC-MS/MS for protein identification. Twelve proteins were verified, of which four were reported involvement of the regulation of pH homeostasis in cytoplasm of alkaliphiles, and the others were originally found their protein expression responding to the environment with different pH values.