RRC ID 30163
Author Kurokawa Y, Yanagi H, Yura T.
Title Overexpression of protein disulfide isomerase DsbC stabilizes multiple-disulfide-bonded recombinant protein produced and transported to the periplasm in Escherichia coli.
Journal Appl. Environ. Microbiol.
Abstract Dsb proteins (DsbA, DsbB, DsbC, and DsbD) catalyze formation and isomerization of protein disulfide bonds in the periplasm of Escherichia coli. By using a set of Dsb coexpression plasmids constructed recently, we analyzed the effects of Dsb overexpression on production of horseradish peroxidase (HRP) isozyme C that contains complex disulfide bonds and tends to aggregate when produced in E. coli. When transported to the periplasm, HRP was unstable but was markedly stabilized upon simultaneous overexpression of the set of Dsb proteins (DsbABCD). Whereas total HRP production increased severalfold upon overexpression of at least disulfide-bonded isomerase DsbC, maximum transport of HRP to the periplasm seemed to require overexpression of all DsbABCD proteins, suggesting that excess Dsb proteins exert synergistic effects in assisting folding and transport of HRP. Periplasmic production of HRP also increased when calcium, thought to play an essential role in folding of nascent HRP polypeptide, was added to the medium with or without Dsb overexpression. These results suggest that Dsb proteins and calcium play distinct roles in periplasmic production of HRP, presumably through facilitating correct folding. The present Dsb expression plasmids should be useful in assessing and dissecting periplasmic production of proteins that contain multiple disulfide bonds in E. coli.
Volume 66(9)
Pages 3960-5
Published 2000-9
PMID 10966415
PMC PMC92245
MeSH Calcium Chloride / pharmacology Disulfides / chemistry Escherichia coli / enzymology* Escherichia coli / genetics Escherichia coli / growth & development Horseradish Peroxidase / genetics Horseradish Peroxidase / metabolism* Periplasm / enzymology* Periplasm / metabolism Protein Disulfide-Isomerases / genetics Protein Disulfide-Isomerases / metabolism* Protein Folding Protein Transport Recombinant Proteins / genetics Recombinant Proteins / metabolism
IF 3.807
Times Cited 45
Prokaryotes E. coli ME7982 ME7983