Thi20, a remarkable enzyme from Saccharomyces cerevisiae with dual thiamin biosynthetic and degradation activities.

Saccharomyces cerevisiae Thi20 is a fusion protein with homology to Bacillus subtilis ThiD and TenA. The N-terminus of Thi20 has significant sequence homology to B. subtilis ThiD, while the C-terminus has homology to B. subtilis TenA. Incubation of Thi20 with thiamin reveals that it has thiaminase II activity, in addition, incubation of Thi20 with HMP (4-amino-2-methyl-5-hydroxymethylpyrimidine) and ATP reveals that it has HMP kinase and HMP-P (4-amino-2-methyl-5-hydroxymethylpyrimidine phosphate) kinase activity. This demonstrates that Thi20 is a trifunctional protein with thiamin biosynthetic and degradative activity.

Structural characterization of the regulatory proteins TenA and TenI from Bacillus subtilis and identification of TenA as a thiaminase II.

Bacillus subtilis gene products TenA and TenI have been implicated in regulating the production of extracellular proteases, but their role in the regulation process remains unclear. The structural characterization of these proteins was undertaken to help provide insight into their function. We have determined the structure of TenA alone and in complex with 4-amino-2-methyl-5-hydroxymethylpyrimidine, and we demonstrate that TenA is a thiaminase II. The TenA structure suggests that the degradation of thiamin by TenA likely proceeds via the same addition-elimination mechanism described for thiaminase I. Three active-site residues, Asp44, Cys135, and Glu205, are likely involved in substrate binding and catalysis based on the enzyme/product complex structure and the conservation of these residues within TenA sequences. We have also determined the structure of TenI. Although TenI shows significant structural homology to thiamin phosphate synthase, it has no known enzymatic function. The structure suggests that TenI is unable to bind thiamin phosphate, largely resulting from the presence of leucine at position 119, while the corresponding residue in thiamin phosphate synthase is glycine.