Huang Yu-Ju, Tsai Tsung-Yu, Pan Tzu-Ming
Institute of Microbiology and Biochemistry, National Taiwan University, Taipei, Taiwan, Republic of China.
J Agric Food Chem. 2007 Aug 22;55(17):7182-91. doi: 10.1021/jf071014s. Epub 2007 Jul 14.
Escherichia coli O157:H7 has an unusually high resistance to acidic environments. Some research has revealed that acid-adapted cells, by exposure to moderately acidic conditions, are more resistant to a subsequent strong acidic challenge or other stress. This study was conducted to understand the protein expression regulation of acid tolerance response (ATR) of a local isolated E. coli O157:H7 TWC01 (TWC01) induced by an acidic environment. TWC01 cells were acid adapted by using hydrochloric acid (HCl) or lactic acid as acidifier to induce ATR. The total proteins of adapted cells were extracted for proteomic analysis and protein identification by matrix-assisted laser desorption ionization quadrupole time-of-flight tandem mass spectrometry (MALDI-Q-TOF MS/MS). Furthermore, the effects of acid adaptation on shiga-like toxin (stx) secretion were examined. Results revealed that acid adaptation depressed stx production of E. coli O157:H7 TWC01 during adaptation and did not improve post-stress toxin production. Image analysis of the gel indicated that numerous proteins were up-regulated and that lactic acid had a greater effect than HCl did (percentages of up-regulated proteins were 57.64 and 35.47%, respectively). Analysis of proteins by mass spectrometry revealed that most of the up-regulated proteins were metabolism-related, including phosphoglycerate kinase (PGK), glutamate decarboxylases alpha and beta (GadA, GadB), adenine phosphoribosyltransferase (APRT), and dihydrodipicolinate synthase (DHDPS). Others were related to translation (e.g., elongation factor Tu, elongation factor G), protein folding (e.g., alkyl hydroperoxide reductase), and membrane proteins (e.g., ompA precursor and ompR). The variation of protein expression showed that acid resistance was induced in TWC01 and was primarily manifested via expression of up-regulated proteins that contribute to increased energy conservation and polypeptide synthesis.
大肠杆菌O157:H7对酸性环境具有异常高的耐受性。一些研究表明，通过暴露于适度酸性条件下而适应酸性环境的细胞，对随后的强酸性挑战或其他应激更具抵抗力。本研究旨在了解酸性环境诱导的本地分离大肠杆菌O157:H7 TWC01（TWC01）的耐酸性反应（ATR）的蛋白质表达调控。通过使用盐酸（HCl）或乳酸作为酸化剂使TWC01细胞适应酸性环境以诱导ATR。提取适应细胞的总蛋白用于蛋白质组学分析，并通过基质辅助激光解吸电离四极杆飞行时间串联质谱（MALDI-Q-TOF MS/MS）进行蛋白质鉴定。此外，还研究了酸性适应对志贺样毒素（stx）分泌的影响。结果显示，酸性适应在适应过程中抑制了大肠杆菌O157:H7 TWC01的stx产生，并且并未提高应激后毒素的产生。凝胶图像分析表明，大量蛋白质上调，且乳酸的作用比HCl更大（上调蛋白质的百分比分别为57.64%和35.47%）。通过质谱分析蛋白质发现，大多数上调的蛋白质与代谢相关，包括磷酸甘油酸激酶（PGK）、谷氨酸脱羧酶α和β（GadA、GadB）、腺嘌呤磷酸核糖转移酶（APRT）和二氢二吡啶二羧酸合酶（DHDPS）。其他蛋白质与翻译（如延伸因子Tu、延伸因子G）、蛋白质折叠（如烷基过氧化氢还原酶）和膜蛋白（如ompA前体和ompR）有关。蛋白质表达的变化表明，TWC01诱导了耐酸性，主要通过上调有助于增加能量保存和多肽合成的蛋白质的表达来体现。
