Purification and some properties of thiamine-binding protein from rice bran.

A thiamine-binding protein was purified from the extract of rice bran acetone powder by conventional procedures of acid precipitation, a series of column chromatography on DEAE-Sephadex A-50 and DEAE-cellulose, and gel filtration of Sephadex G-200. The purified thiamine-binding protein was nearly homogeneous as judged by disc gel electrophoresis and the molecular weight was estimated to be 94,000 by gel filtration on Sephadex Gn-200 and 50,000 by sodium dodecylsulfate (SDS) gel electrophoresis, suggesting that the protein is composed of two identical subunits. The apparent Kd and Bmax of the binding for [14C]thiamine was 0.44 +/- 0.05 microM and 17.2 +/- 0.7 nmol/mg of protein, respectively. The optimal pH for the binding is between 8.0 and 9.0. From the competition experiment using several thiamine derivatives, high binding specificity of the protein for thiamine was presumed.