title: Effect of oxidized and reduced forms of Escherichia coli DsbC on protein refolding.
abstract: DsbC, which catalyzes disulfide isomerization, was overproduced in the periplasm of Escherichia coli and purified from the periplasmic fraction by osmotic shock and anion-exchange chromatography. The active site of the purified DsbC was found to be an oxidized form (ox-DsbC) which could be converted to the reduced form (red-DsbC) by the addition of dithiothreitol. The effect of ox- and red-DsbC on the refolding of chemically denatured and reduced proteins with different numbers of disulfide bonds and free cysteine-thiol groups was investigated. Ox-DsbC facilitated the refolding of proteins with multiple disulfide bonds in both oxidative and reductive environments, while red-DsbC facilitated refolding only in the former. On the other hand, only red-DsbC facilitated the refolding of proteins with multiple free cysteine-thiol groups but either form of DsbC did not facilitate the refolding of proteins with only one cysteine-thiol group. It is therefore important to choose the form which suits the properties of the protein. Holo-chaperonin from Thermus thermophilus and DsbC demonstrated a synergistic effect on protein refolding.
authors.affiliation: Division of Molecular Science, Graduate School of Science and Technology, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe 657-8501, Japan.
authors.forename: Jiro
authors.initials: J
authors.lastname: Kohda
authors.forename: Noboru
authors.initials: N
authors.lastname: Kawahara
authors.forename: Hideki
authors.initials: H
authors.lastname: Fukuda
authors.forename: Akihiko
authors.initials: A
authors.lastname: Kondo