Isolation and Characterization of Calf Liver Thioredoxin

Abstract

Abstract The reduced form of thioredoxin, a low molecular weight hydrogen transport protein, was isolated from calf liver. The purification involved heat treatment and chromatography on DEAE-cellulose, Sephadex G-50, and CM-cellulose, and it resulted in an apparently pure protein after a 4,000-fold purification with a final yield of 30%. The thioredoxin preparation obtained was homogeneous as judged by polyacrylamide gel electrophoresis and the presence of valine as the only NH2-terminal amino acid. Chromatography on Sephadex indicated a molecular weight of approximately 12,000 for liver thioredoxin and amino acid analysis showed a content of 103 amino acid residues, including 4 half-cystine residues. Oxidation in vitro of the reduced form of calf liver thioredoxin indicated that only 2 of the half-cystine residues formed an oxidation-reduction-active disulfide bridge.