Department of Chemical and Biological Engineering, Korea University, Anam-Dong 5-1, Seongbuk-Gu, Seoul 136-713, Republic of Korea.
Enzyme Microb Technol. 2011 Jul 10;49(2):124-30. doi: 10.1016/j.enzmictec.2011.04.013. Epub 2011 Apr 22.
Through the proteome analysis of Escherichia coli BL21(DE3), we previously identified the stress-responsive protein, arsenate reductase (ArsC), that showed a high cytoplasmic solubility and a folding capacity even in the presence of stress-inducing reagents. In this study, we used ArsC as an N-terminal fusion partner to synthesize nine aggregation-prone proteins as water-soluble forms. As a result, solubility of the aggregation-prone proteins increased dramatically by the fusion of ArsC, due presumably to its tendency to facilitate the folding of target proteins. Also, we evaluated and confirmed the efficacy of ArsC-fusion expression in making the fusion-expressed target proteins have their own native function or structure. That is, the self-assembly function of human ferritin light chain, l-arginine-degrading function of arginine deiminase, and the correct secondary structure of human granulocyte colony stimulating factor were clearly observed through transmission electron microscope analysis, colorimetric enzyme activity assay, and circular dichroism, respectively. It is strongly suggested that ArsC can be in general an efficient fusion expression partner for the production of soluble and active heterologous proteins in E. coli.
通过对大肠杆菌 BL21(DE3)的蛋白质组分析，我们先前鉴定出应激响应蛋白砷酸盐还原酶（ArsC），即使在存在应激诱导试剂的情况下，它也表现出高细胞质可溶性和折叠能力。在这项研究中，我们将 ArsC 用作 N 端融合伙伴，以水溶性形式合成了 9 种易聚集的蛋白质。结果，由于 ArsC 促进靶蛋白折叠的趋势，易聚集蛋白的可溶性显著增加。此外，我们还评估并证实了 ArsC 融合表达在使融合表达的靶蛋白具有其自身天然功能或结构方面的功效。也就是说，通过透射电子显微镜分析、比色酶活性测定和圆二色性，分别清楚地观察到了人铁蛋白轻链的自组装功能、精氨酸脱氨酶的 l-精氨酸降解功能以及人粒细胞集落刺激因子的正确二级结构。强烈建议 ArsC 通常可以作为大肠杆菌中生产可溶性和活性异源蛋白的有效融合表达伙伴。
