Engineered DsbC chimeras catalyze both protein oxidation and disulfide-bond isomerization in Escherichia coli: Reconciling two competing pathways.
Engineered DsbC chimeras catalyze both protein oxidation and disulfide-bond isomerization in Escherichia coli: Reconciling two competing pathways.
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工程化 DsbC 嵌合体催化大肠杆菌中的蛋白质氧化和二硫键异构化：协调两条竞争途径。
DOI：
10.1073/pnas.0403003101
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发表时间：
2004
期刊：
影响因子：
--
通讯作者：
Georgiou,George
中科院分区：
文献类型：
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作者：
Segatori,Laura;Paukstelis,PaulJ;Gilbert,HiramF;Georgiou,George
In theEscherichia coliperiplasm, the formation of protein disulfide bonds is catalyzed by DsbA and DsbC. DsbA is a monomer that is maintained in a fully oxidized state by the membrane enzyme DsbB, whereas DsbC is a dimer that is kept reduced by a second membrane protein, DsbD. Although the catalytic regions of DsbA and DsbC are composed of structurally homologous thioredoxin motif domains, DsbA serves only as an oxidasein vivo, whereas DsbC catalyzes disulfide reduction and isomerization and also exhibits significant chaperone activity. To reconcile the distinct catalytic activities of DsbC and DsbA, we constructed a series of chimeras comprising of the dimerization domain of DsbC, with or without the adjacent α-helical linker region, fused either to the first, second, third, or fifth residue of intact DsbA or to thioredoxin. The chimeras fully substituted for DsbC in disulfide-bond rearrangement and also were able to restore protein oxidation in adsbAbackground. Remarkably, the chimeras could serve as a single catalyst for both disulfide-bond formation and rearrangement, thus reconciling the kinetically competing DsbB–DsbA and DsbD–DsbC pathways. This property appeared to depend on the orientation of the DsbA active-site cysteines with respect to the DsbC dimerization domain.In vitro, the chimeras had high chaperone activity and significant reductase activity but only 15–22% of the disulfide-isomerization activity of DsbC, suggesting that rear-rangement of nonnative disulfides may be mediated primarily by cycles of random reduction and reoxidation.
