CHARACTERIZATION OF A NOVEL BI-FUNCTIONAL DIHYDROPTEROATE SYNTHASE/DIHYDROPTEROATE REDUCTASE ENZYME FROM Helicobacter pylori

The Department of Molecular Microbiology and Biotechnology, George S. Wise Faculty of Life Sciences, Tel-Aviv University, Tel-Aviv 69978, Israel; Department of Biological Chemistry, University of Michigan, Ann-Arbor Michigan MI 48109-0606, USA

^* To whom correspondence should be addressed. Email: mevarech{at}post.tau.ac.il.

	Abstract

Tetrahydrofolate is a ubiquitous C1 carrier in many biosynthetic pathways in bacteria, importantly in the biosynthesis of formyl-methionyl tRNA^fMet which is essential for initiation of translation. The final step in the biosynthesis of tetrahydrofolate is carried out by the enzyme dihydrofolate reductase (DHFR). A search of the complete genome sequence of Helicobacter pylori failed to reveal any sequence that encodes for DHFR. Previous studies demonstrated that the H. pylori dihydropteroate synthase gene folP can complement an Escherichia coli strain in which folA and folM, encoding two distinct DHFRs, were deleted. It was also shown that the H. pylori FolP possesses an additional N-terminal domain that binds FMN. Homologous domains are found in FolP of other microorganisms that do not possess DHFR. In this communication we demonstrate that the H. pylori FolP is also a dihydropteroate reductase that derives its reducing power from soluble flavins, FMNH₂ and FADH₂. We also determined the stoichiometry of the enzyme bound flavin and showed that half of the bound flavin is exchangeable with the soluble flavins. Finally, site directed mutagenesis of the most conserved amino acid residues in the N-terminal domain indicated the importance of these residues for the activity of the enzyme as dihydropteroate reductase.