Gadgil Mugdha, Kapur Vivek, Hu Wei-Shou
Department of Chemical Engineering and Materials Science, Biomedical Genomics Center, University of Minnesota, 421 Washington Avenue SE, Minneapolis, Minnesota 55455-0132, USA.
Biotechnol Prog. 2005 May-Jun;21(3):689-99. doi: 10.1021/bp049630l.
Temperature shift is often practiced in the cultivation of Escherichia coli to reduce undesired metabolite formation and to maximize synthesis of correctly folded heterologous protein. As the culture temperature is decreased below the optimal 37 degrees C, growth rate decreases and many physiological changes occur. In this study, we investigated the gene expression dynamics of E. coli on switching its cultivation temperature from 37 to 33 and 28 degrees C using whole genome DNA microarrays. Approximately 9% of the genome altered expression level on temperature shift. Overall, the alteration of transcription upon the downshift of temperature is rapid and globally distributed over a wide range of gene classes. The general trends of transcriptional changes at 28 and 33 degrees C were similar. The largest functional class among the differentially expressed genes was energy metabolism. About 12% of genes in energy metabolism show a decrease in their level of expression, and approximately 6% show an increase. Consistent with the decrease in the glucose uptake rate, many genes involved in glycolysis and the PTS sugar transport systems show decreased expression. Genes encoding enzymes related to amino acid biosynthesis and transport also have reduced expression levels. Such decrease in expression probably reflects the reduced growth rate and the accompanying reduction in energy and amino acid demand at lower temperatures. However, nearly all genes encoding enzymes in the TCA cycle have increased expression levels, which may well be compensating the reduction of the activity of TCA cycle enzymes at lower temperatures. Temperature shift also results in shift of the cytochromes from the high affinity cytochrome o system to the low affinity cytochrome d system. There is no evidence that protein processing genes are selectively altered to create favorable conditions for heterologous protein synthesis. Our results indicate that the beneficial effect of temperature shift in many biotechnological processes is likely to be attributed to the general effect of reduced growth and metabolism.
温度转换常用于大肠杆菌的培养，以减少不期望的代谢物形成，并使正确折叠的异源蛋白的合成最大化。当培养温度降至最佳的37℃以下时，生长速率下降，并发生许多生理变化。在本研究中，我们使用全基因组DNA微阵列研究了大肠杆菌在将培养温度从37℃切换到33℃和28℃时的基因表达动态。约9%的基因组在温度转换时改变了表达水平。总体而言，温度下降时转录的改变迅速且广泛分布于多种基因类别。28℃和33℃时转录变化的总体趋势相似。差异表达基因中最大的功能类别是能量代谢。能量代谢中约12%的基因表达水平下降，约6%的基因表达水平上升。与葡萄糖摄取率的降低一致，许多参与糖酵解和磷酸转移酶系统糖转运的基因表达下降。编码与氨基酸生物合成和转运相关酶的基因表达水平也降低。这种表达下降可能反映了较低温度下生长速率的降低以及随之而来的能量和氨基酸需求的减少。然而，几乎所有编码三羧酸循环中酶的基因表达水平都有所增加，这很可能是在补偿较低温度下三羧酸循环酶活性的降低。温度转换还导致细胞色素从高亲和力的细胞色素o系统转变为低亲和力的细胞色素d系统。没有证据表明蛋白质加工基因被选择性改变以创造有利于异源蛋白合成的条件。我们的结果表明，温度转换在许多生物技术过程中的有益作用可能归因于生长和代谢降低的总体效应。
