Hoshino Kyoko, Eda Akiko, Kurokawa Yoichi, Shimizu Norio
Faculty of Life Sciences, Toyo University, Ora-gun, Gunma, Japan.
Biosci Biotechnol Biochem. 2002 Feb;66(2):344-50. doi: 10.1271/bbb.66.344.
When brain-derived neurotrophic factor (BDNF) is produced in the Escherichia coli periplasm, insoluble BDNF proteins with low biological activity and having mismatched disulfide linkages are formed. The coexpression of cysteine oxidoreductases (DsbA and DsbC) and membrane-bound enzymes (DsbB and DsbD), which play an important role in the formation of disulfide bonds in the periplasm, was investigated to improve the production of soluble and biologically active BDNF. The expression levels of Dsb proteins changed when the growth medium and the Dsb expression plasmids were changed, and the production rate of soluble BDNF was almost proportional to the expression level of DsbC protein with disulfide isomerase activity in the case of a low expression level of BDNF. The rate of soluble BDNF production with coexpression of DsbABCD was as high as 35%. These results show that coexpression of BDNF and Dsb proteins can effectively increase the production of soluble and biologically active BDNF.
当在大肠杆菌周质中产生脑源性神经营养因子（BDNF）时，会形成具有低生物活性且二硫键错配的不溶性BDNF蛋白。为了提高可溶性且具有生物活性的BDNF的产量，研究了在周质中二硫键形成过程中起重要作用的半胱氨酸氧化还原酶（DsbA和DsbC）以及膜结合酶（DsbB和DsbD）的共表达情况。当生长培养基和Dsb表达质粒发生变化时，Dsb蛋白的表达水平也会改变，并且在BDNF低表达水平的情况下，可溶性BDNF的产生率几乎与具有二硫键异构酶活性的DsbC蛋白的表达水平成正比。共表达DsbABCD时可溶性BDNF的产生率高达35%。这些结果表明，BDNF与Dsb蛋白的共表达可以有效地提高可溶性且具有生物活性的BDNF的产量。
