Pleitner Aaron, Zhai Yong, Winter Roland, Ruan Lifang, McMullen Lynn M, Gänzle Michael G
University of Alberta, Department of Agricultural, Food and Nutritional Science, Edmonton, Canada T6G 2P5.
Biochim Biophys Acta. 2012 Dec;1824(12):1351-7. doi: 10.1016/j.bbapap.2012.07.007. Epub 2012 Jul 27.
This study investigated the mechanisms of heat resistance in Escherichia coli AW1.7 by quantification of cytoplasmic solutes, determination of ribosome denaturation, and by determination of protein denaturation. To assess the contribution of heat shock proteins and compatible solutes, experiments were conducted after exposure to sublethal heat shock, and with cultures grown at NaCl concentrations ranging from 0 to 6%. Heat resistance of E. coli AW1.7 was compared to the heat sensitive E. coli GGG10 and a plasmid-cured, heat sensitive derivative of E. coli AW1.7 named E. coli AW1.7ΔpHR1. Sublethal heat shock improved survival at 60°C of E. coli GGG10 and AW1.7ΔpHR1 but not of E. coli AW1.7. Addition of NaCl increased the heat resistance of all three strains, but only E. coli AW1.7 exhibited high heat resistance when grown in NaCl concentrations ranging from 2 to 6%. E. coli AW1.7 and GGG10 accumulated 16.1 ± 0.8 and 8.8 ± 0.8mmolL⁻¹ amino acids when grown at 0% NaCl, and 1.47 ± 0.07 and 0.78 ± 0.06mmolL⁻¹ carbohydrates when grown at 6% NaCl, respectively. Ribosome denaturation was determined by differential scanning calorimetry. After growth in the presence of 0% NaCl, the 30S subunit denatured at 63.7 ± 0.8°C and 60.7 ± 0.3°C in E. coli AW1.7 and GGG10, respectively. Fourier-transformed-infrared-spectroscopy did not indicate differences in protein denaturation between the strains during heating. In conclusion, heat resistance in E. coli AW1.7 correlates to ribosome stability at 60°C and is dependent on accumulation of cytoplasmic solutes.
本研究通过对细胞质溶质进行定量分析、测定核糖体变性以及蛋白质变性，探究了大肠杆菌AW1.7的耐热机制。为评估热休克蛋白和相容性溶质的作用，在亚致死热休克处理后以及在NaCl浓度为0%至6%的条件下培养细菌时进行了实验。将大肠杆菌AW1.7的耐热性与热敏性大肠杆菌GGG10以及大肠杆菌AW1.7的质粒治愈型热敏衍生物（命名为大肠杆菌AW1.7ΔpHR1）进行了比较。亚致死热休克提高了大肠杆菌GGG10和AW1.7ΔpHR1在60°C时的存活率，但对大肠杆菌AW1.7没有作用。添加NaCl提高了所有三种菌株的耐热性，但只有大肠杆菌AW1.7在2%至6%的NaCl浓度下生长时表现出高耐热性。当在0% NaCl条件下生长时，大肠杆菌AW1.7和GGG10分别积累了16.1±0.8和8.8±0.8 mmol·L⁻¹的氨基酸，当在6% NaCl条件下生长时，分别积累了1.47±0.07和0.78±0.06 mmol·L⁻¹的碳水化合物。通过差示扫描量热法测定核糖体变性。在0% NaCl存在下生长后，大肠杆菌AW1.7和GGG10中30S亚基的变性温度分别为63.7±0.8°C和60.7±0.3°C。傅里叶变换红外光谱法未显示加热过程中各菌株之间蛋白质变性存在差异。总之，大肠杆菌AW1.7的耐热性与60°C时核糖体的稳定性相关，并且依赖于细胞质溶质的积累。
