STRUCTURAL STUDIES ON FPGS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Folic acid and its derivatives are essential vitamins required for cell growth and replication in both prokaryotic and eukaryotic cells, primarily through their central role in a variety of reactions involved in one-carbon metabolism. Folic acid serves as a carrier of single carbon units typically in the form of methyl and formyl substituents, and these folate derivatives are involved in a number of diverse biochemical pathways, including methionine metabolism, the biosynthesis of purine nucleotides and thymidylate synthesis. Folates within the cell are generally found as polyglutamate derivatives, and it has been shown that many folate-dependent enzymes have a higher affinity for the polyglutamylated forms, relative to the monoglutamate. In organisms that synthesize folates de novo (eg. plants, bacteria, fungi and protozoa), two enzymes catalyze the addition of glutamic acid residues; firstly dihydrofolate synthetase (DHFS) adds the first glutamic acid residue to dihyropteroate to produce dihydrofrolate (DHF). The ubiquitous enzyme folylpolyglutamate synthetase (FPGS, EC 6.3.2.17) then catalyzes folate polyglutamylation in all cells including those that require exogenous folate (eg. mammalian cells). In some bacteria DHFS and FPGS activity resides on a single gene (eg. Escherichia coli and Corynebacterium), whereas in plants and fungi it appears that two distinct genes encode for the two enzymes. The crystal structure of FPGS from three bacterial species have been determined from Lactobacillus casei, Thermotoga maritima and the bifunctional DHFS/FPGS E. coli. We have recently solved the structures of Mycobacterium tuberculosis FPGS in complex with ADP and AMPPCP, and three site-directed mutants of L. casei FPGS.