Isolation and characterization of thiamin-binding protein from chicken egg white

Abstract

A thiamin-binding protein was isolated and characterized from chicken egg white by affinity chromatography on thiamin pyrophosphate coupled to aminoethyl-Sepharose. The high specificity of interaction between the thiamin-binding protein and the riboflavin-binding protein of the egg white, with a protein/protein molar ratio of 1.0, led to the development of an alternative procedure that used the riboflavin-binding protein immobilized on CNBr-activated Sepharose as the affinity matrix. The thiamin-binding protein was homogeneous by the criteria of polyacrylamide gel disc electrophoresis, double immunodiffusion and sodium dodecyl sulphate/polyacrylamide gel electrophoresis, had a MW of 38,000 .+-. 2000 and was not a glycoprotein. The protein bound [14C]thiamin with a molar ratio of 1.0, with Kd 0.3 .mu.M.