Hydroxyethylthiazole uptake in Escherichia coli: general properties and relationship between uptake and thiamine...

Uptake of (35)S-hydroxyethylthiazole (4-methyl-5-hydroxyethylthiazole) by Escherichia coli intact cells was studied. Hydroxyethylthiazole was taken up in the presence and absence of glucose at the same rate. The uptake was almost proportional to a hydroxyethylthiazole concentration gradient up to 0.1 mM with no tendency of saturation, and reached a steady state within 2 min. When the cells were treated with 1 mM N-ethylmaleimide, about 50% inhibition of hydroxyethylthiazole uptake was observed. Hydroxyethylthiazole uptake was stimulated by the addition of hydroxymethylpyrimidine (2-methyl-4-amino-5-hydroxymethylpyrimidine), and this effect was further enhanced in the presence of glucose. For full activation of hydroxyethylthiazole uptake, 1 muM hydroxymethylpyrimidine was necessary in the presence of glucose. The rate of hydroxyethylthiazole uptake was almost linear up to 60 min in the presence of hydroxymethylpyrimidine and glucose. Hydroxymethylpyrimidine monophosphate and its pyrophosphate could not stimulate the uptake. Thiamine and 2-amino-hydroxyethylthiazole were inhibitory on hydroxyethylthiazole uptake in the presence of hydroxymethylpyrimidine and glucose. N-ethylmaleimide and 2, 4-dinitrophenol were also inhibitory. No stimulatory effect of hydroxymethylpyrimidine on hydroxyethylthiazole uptake was observed in mutant cells lacking either thiaminephosphate pyrophosphorylase or hydroxymethylpyrimidine monophosphate kinase. The possibility of direct participation of thiamine-synthesizing enzymes in hydroxyethylthiazole uptake was discussed.