Open AccessRecombinant expression and in vitro folding of proinsulin are stimulated by the synthetic dithiol Vectrase‐P
Author(s) -
Winter Jeannette,
Lilie Hauke,
Rudolph Rainer
Publication year - 2002
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2002.tb11310.x
Subject(s) - proinsulin , periplasmic space , dithiol , biochemistry , protein disulfide isomerase , protein folding , chemistry , folding (dsp implementation) , recombinant dna , isomerase , in vitro , escherichia coli , redox , biology , enzyme , insulin , engineering , organic chemistry , electrical engineering , gene , endocrinology
Recombinant production of native proinsulin in the periplasm of Escherichia coli is limited by formation of the correct disulfide bonds and inclusion body formation. These limitations can be overcome during in vitro folding of proinsulin by using a redox system and also protein disulfide isomerase. Here, we added a redox active substance, Vectrase‐P, to the cultivation medium of E. coli cells producing proinsulin. We show that this synthetic dithiol partially mimicking the redox activity of protein disulfide isomerase provides an improved redox situation in the periplasm and, therefore, provides optimum conditions for folding of proinsulin in that cell compartment resulting in an increase in yield of 60%. The in vivo results were confirmed by analyzing in vitro folding of proinsulin in the presence of the dithiol Vectrase‐P.