[Characteristics of thiamine-binding protein from rat brain synaptosomes]

The thiamine-binding protein was obtained from rat brain synaptosomes by affinity chromatography and gel-filtration on Sephadex G-200. The protein is homogeneous by the data of SDS gel-electrophoresis, anode electrophoresis and isofocusing between pH 3.5-9.0. The isoelectric point of this protein is near pH 4.8-5.0. The binding nature of the protein with [14C] thiamine was studied. It is shown that metal ions, especially Na+ and Ca2+, increase the thiamine-binding activity. The binding process is of a saturation character at the thiamine concentrations of 10(-7)-10(-5) M. Thiamine possesses two binding sites with KD1 = 3.1 microM and KD2 = 30 microM. Out of the tested thiamine analogues and antagonists of vitamin B1 thiamine-monophosphate and pyrithiamine were the most competitive.