Mu Tingzhen, Yang Maohua, Zhao Jixiang, Sharshar Moustafa Mohammed, Tian Jiangnan, Xing Jianmin
State Key Laboratory of Biochemical Engineering and Key Laboratory of Green Process and Engineering, Institute of Process Engineering, Chinese Academy of Sciences, P.O. Box 353, Beijing, 100190, People's Republic of China.
University of Chinese Academy of Sciences, Beijing, 100049, People's Republic of China.
Biotechnol Lett. 2017 Mar;39(3):447-452. doi: 10.1007/s10529-016-2266-1. Epub 2016 Dec 20.
To construct efficient transformation and expression system and further improve desulfurizing activity of cells through expression of Vitreoscilla hemoglobin (VHb) in haloalkaliphilic Thialkalivibrio versutus SOB306.
We transferred plasmids pKT230 and pBBR-sm into T. versutus SOB306 via a conjugation method. We identified four promoters from among several predicted promoters by scoring for streptomycin resistance, and finally selected tac and p3 based on the efficiency of expression of red fluorescent protein (RFP). Expression of RFP when regulated by tac was more than three times that of p3 in SOB306. Further, we expressed VHb under the control of tac promoter in SOB306. Expression of VHb was verified using CO-difference spectra. The results showed that VHb expression can boost sulfur metabolism, as evidenced by an increase of about 11.7 ± 1.8% in the average rate of thiosulfate removal in the presence of VHb.
A conjugation transfer and an expression system for Thialkalivibrio, has been developed for the first time and used for expression of VHb to improve desulfurizing activity.
构建高效的转化和表达系统，并通过在嗜盐碱多硫代碱弧菌SOB306中表达透明颤菌血红蛋白（VHb）进一步提高细胞的脱硫活性。
我们通过接合方法将质粒pKT230和pBBR-sm转入多硫代碱弧菌SOB306。我们通过对链霉素抗性进行评分，从几个预测的启动子中鉴定出四个启动子，最后根据红色荧光蛋白（RFP）的表达效率选择了tac和p3。在SOB306中，由tac调控时RFP的表达是p3调控时的三倍多。此外，我们在tac启动子的控制下在SOB306中表达了VHb。使用CO差光谱验证了VHb的表达。结果表明，VHb的表达可以促进硫代谢，在存在VHb的情况下，硫代硫酸盐去除平均速率提高约11.7±1.8%，这证明了这一点。
首次开发了一种用于多硫代碱弧菌的接合转移和表达系统，并用于表达VHb以提高脱硫活性。
