突变体
蛋白酶
信号肽
生物化学
蛋白质二硫键异构酶
生物
分泌物
互补DNA
大肠杆菌
分泌蛋白
异构酶
分子生物学
酶
重组DNA
化学
基因
作者
Toshitaka Kajino,Kiyoko Kato,Chikara Miyazaki,Osamu Asami,Maretoshi Hirai,Yukio Yamada,Shigezo Udaka
标识
DOI:10.1016/s1389-1723(99)80005-x
摘要
The efficient production of a thermostable protein disulfide isomerase (PDI) was successfully achieved using the newly isolated protease-deficient mutant, Bacillus brevis 31-OK. Extracellular protease (exoprotease) activity was about a quarter of that in the parent, and the mutant was deficient in at least one of the major exoproteases. The cDNA encoding the fungal PDI was inserted downstream of the signal peptide-encoding region in an expression-secretion vector for B. brevis. Efficient production of PDI was feasible using B. brevis 31-OK as a host and modified signal sequences composed of three leucine residues inserted in the hydrophobic region of the MWP (middle wall protein) signal sequence. The maximal secretion of PDI into the culture medium was 1.1 g/l, which is about twice that by the parent strain and fifty times greater than the amount of rat and murine PDIs produced by Escherichia coli. The enzymatic properties such as the specific activity and thermal stability of the recombinant PDI are similar to those of natural PDI derived from Humicola insolens mycelia. B. brevis 31-OK was able to maintain its exoprotease activity at a low level throughout the cultivation and is considered to be useful host for production of a protease-sensitive protein and for increase of protein productivity due to stable accumulation.
科研通智能强力驱动
Strongly Powered by AbleSci AI