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Functional in vitro analysis of the ERO1 protein and protein-disulfide isomerase pathway.

ABSTRACT: Oxidative protein folding in the endoplasmic reticulum is supported by efficient electron relays driven by enzymatic reactions centering on the ERO1-protein-disulfide isomerase (PDI) pathway. A controlled in vitro oxygen consumption assay was carried out to analyze the ERO1-PDI reaction. The results showed the pH-dependent oxidation of PDI by ERO1?. Among several possible disulfide bonds regulating ERO1? activity, Cys(94)-Cys(131) and Cys(99)-Cys(104) disulfide bonds are dominant regulators by excluding the involvement of the Cys(85)-Cys(391) disulfide in the regulation. The fine-tuned species specificity of the ERO1-PDI pathway was demonstrated by functional in vitro complementation assays using yeast and mammalian oxidoreductases. Finally, the results provide experimental evidence for the intramolecular electron transfer from the a domain to the a' domain within PDI during its oxidation by ERO1?.

SUBMITTER: Araki K

PROVIDER: S-EPMC3173198 | biostudies-other | 2011 Sep

REPOSITORIES: biostudies-other

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Functional in vitro analysis of the ERO1 protein and protein-disulfide isomerase pathway.

Araki Kazutaka K Nagata Kazuhiro K

The Journal of biological chemistry 20110708 37

Oxidative protein folding in the endoplasmic reticulum is supported by efficient electron relays driven by enzymatic reactions centering on the ERO1-protein-disulfide isomerase (PDI) pathway. A controlled in vitro oxygen consumption assay was carried out to analyze the ERO1-PDI reaction. The results showed the pH-dependent oxidation of PDI by ERO1α. Among several possible disulfide bonds regulating ERO1α activity, Cys(94)-Cys(131) and Cys(99)-Cys(104) disulfide bonds are dominant regulators by e ...[more]

PMID: 21757736

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