Structural and mutational analysis of TenA protein (HP1287) from the <i>Helicobacter pylori</i> thiamin salvage pathway – evidence of a different substrate specificity

Abstract

HP1287 (tenA) from Helicobacter pylori is included among the genes that play a relevant role in bacterium colonization and persistence. The gene has been cloned and its product, protein TenA, has been expressed and purified. The crystal structures of the wild-type protein and the mutant F47Y have been determined at resolutions of 2.7 and 2.4 A, respectively. The molecular model, a homotetramer with 222 symmetry, shows that the H. pylori TenA structure belongs to the thiaminase II class of proteins. These enzymes were recently found to be involved in a salvage pathway for the synthesis of the thiamin precursor hydroxypyrimidine, which constitutes a building block in thiamin biosynthesis, in particular in bacteria living in the soil. By contrast, enzymatic measurements on TenA from H. pylori indicate that the activity on the putative substrate 4-amino-5-aminomethyl-2-methylpyrimidine is very modest. Moreover, in the present study, we demonstrate that the mutation at residue 47, a position where a phenylalanine occurs in all the strains of H. pylori sequenced to date, is not sufficient to explain the very low catalytic activity toward the expected substrate. As a result of differences in the colonization environment of H. pylori as well as the TenA structural and catalytic peculiar features, we suggest a possible pivotal role for the H. pylori enzyme in the thiamin biosynthetic route, which is in agreement with the relevance of this protein in the stomach colonization process.