Covalent association of protein disulfide isomerase with recombinant human interleukin 2 in vitro

Protein disulfide isomerase has broad specificity in the catalysis of the formation and rearrangement of native disulfide bonds in proteins. This enzyme has two independent thioredoxin-like active sites (-CGHC-) and a peptide binding site. However, the mechanisms involving the catalytic processes ar...

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Bibliographic Details
Published in:Biochemical and biophysical research communications 1996-06, Vol.223 (1), p.153-159
Main Authors: Ye, J M, Key, C J, Wolfe, J L
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Animals
Antibodies, Monoclonal
Binding Sites
Blotting, Western
Electrophoresis, Polyacrylamide Gel
Humans
Interleukin-2 - isolation & purification
Interleukin-2 - metabolism
Isomerases - chemistry
Isomerases - isolation & purification
Isomerases - metabolism
Mice
Molecular Sequence Data
Oxidation-Reduction
Protein Binding
Protein Disulfide-Isomerases
Recombinant Proteins - isolation & purification
Recombinant Proteins - metabolism
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Summary:Protein disulfide isomerase has broad specificity in the catalysis of the formation and rearrangement of native disulfide bonds in proteins. This enzyme has two independent thioredoxin-like active sites (-CGHC-) and a peptide binding site. However, the mechanisms involving the catalytic processes are not clearly understood. It was reported that the enzyme associates with scrambled pancreatic ribonuclease A in vitro, and with misfolded human lysozyme in vivo. In the present study, recombinant human interleukin 2 has been chosen to probe the reaction intermediate in the reaction with the enzyme. We have identified and characterized a covalent associate formed in vitro by SDS-PAGE and Western blot analysis. This associate has a molecular weight of 71-72 kDa, the approximate sum of the molecular weights of the enzyme and the substrate. Western blot analysis confirmed that it formed via an intermolecular disulfide bond. Upon treatment with 2-mercaptoethanol, this bond was cleaved.
ISSN:0006-291X
DOI:10.1006/bbrc.1996.0861