Snapshots of DsbA in action: detection of proteins in the process of oxidative folding.
DsbA, a thioredoxin superfamily member, introduces disulfide bonds into newly translocated proteins. This process is thought to occur via formation of mixed disulfide complexes between DsbA and its substrates. However, these complexes are difficult to detect, probably because of their short-lived nature. Here we show that it is possible to detect such covalent intermediates in vivo by a mutation in DsbA that alters cis proline-151. Further, this mutant allowed us to identify substrates of DsbA. Alteration of the cis proline, highly conserved among thioredoxin superfamily members, may be useful for the detection of substrates and intermediate complexes in other systems.
Kadokura
Hiroshi
H
Department of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115, USA.
Tian
Hongping
H
Zander
Thomas
T
Bardwell
James C A
JC
Beckwith
Jon
J
D020816:Amino Acid Motifs; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D007536:Isomerism; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008970:Molecular Weight; D009154:Mutation; D010084:Oxidation-Reduction; D010957:Plasmids; D011392:Proline / Q000737:chemistry; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017510:Protein Folding; D013879:Thioredoxins / Q000737:chemistry / Q000378:metabolism; D014161:Transduction, Genetic
D001426:Bacterial Proteins; D004220:Disulfides; C079503:DsbB protein, Bacteria; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D013879:Thioredoxins; D011392:Proline; D019704:Protein Disulfide-Isomerases