Premium
Redox states of DsbA in the periplasm of Escherichia coli
Author(s) -
Kishigami Satoshi,
Akiyama Yoshinori,
Ito Koreaki
Publication year - 1995
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(95)00354-c
Subject(s) - dsba , periplasmic space , chemistry , redox , biochemistry , escherichia coli , inorganic chemistry , gene
DsbA is a periplasmic, disulfide bond formation factor of E. coli . We studied in vivo redox states of its active site cysteines. When periplasmic contents were prepared from iodoacetic acid‐treated cells, according to the previously published procedures, variable but major proportions of DsbA were in the reduced form. We found that this was due to an artificial reduction that occurred after cell disruption; even purified and oxidized DsbA underwent reduction when incubated with cell extracts in the absence of any added reducing agent. Such DsbA‐reducing activities were detected in both the periplasmic and the cytoplasmic fractions. To circumvent the artifact, we analyzed redox states of DsbA under denaturing conditions. Now virtually all the DsbA molecules were detected as oxidized or reduced in the dsbB + background or in the dsbB − background, respectively. Using the improved method, we also examined redox states of DsbA when it was overproduced, and followed the oxidation/reduction pathway that DsbA follows after biosynthesis. It is suggested that newly synthesized DsbA is rapidly oxidized by pre‐existing DsbA, while oxidation of mature (functional) DsbA requires DsbB, whose roles might include that of antagonizing the actions of DsbA‐reducing enzyme(s).