Crystal structures of thymidylate synthase mutant R166Q: structural basis for the nearly complete loss of catalytic activity

Journal of Biochemical and Molecular Toxicology
Rogerio R Sotelo-MundoW R Montfort

Abstract

Thymidylate synthase (TS) catalyzes the folate-dependent methylation of deoxyuridine monophosphate (dUMP) to form thymidine monophosphate (dTMP). We have investigated the role of invariant arginine 166, one of four arginines that contact the dUMP phosphate, using site-directed mutagenesis, X-ray crystallography, and TS from Escherichia coli. The R166Q mutant was crystallized in the presence of dUMP and a structure determined to 2.9 A resolution, but neither the ligand nor the sulfate from the crystallization buffer was found in the active site. A second structure determined with crystals prepared in the presence of dUMP and the antifolate 10-propargyl-5,8-dideazafolate revealed that the inhibitor was bound in an extended, nonproductive conformation, partially occupying the nucleotide-binding site. A sulfate ion, rather than dUMP, was found in the nucleotide phosphate-binding site. Previous studies have shown that the substitution at three of the four arginines of the dUMP phosphate-binding site is permissive; however; for Arg166, all the mutations lead to a near-inactive mutant. The present structures of TS R166Q reveal that the phosphate-binding site is largely intact, but with a substantially reduced affinity for phosphate, d...Continue Reading

References

May 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·M L MichaelsJ H Miller
Jan 1, 1995·Annual Review of Biochemistry·C W Carreras, D V Santi
Jan 7, 1998·Protein Science : a Publication of the Protein Society·P StropW R Montfort
Apr 16, 1998·Pharmacology & Therapeutics·W R Montfort, A Weichsel
Aug 4, 1998·Structure·T J StoutR M Stroud
Jun 30, 2000·Protein Expression and Purification·L M ChangchienF Maley
Apr 24, 2001·Protein Science : a Publication of the Protein Society·R AlmogP Van Roey
Aug 24, 2002·Proceedings of the National Academy of Sciences of the United States of America·Scott A LesleyRaymond C Stevens
Jan 15, 2003·Biochemistry·Robert M Stroud, Janet S Finer-Moore

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Citations

Jun 21, 2014·Journal of the American Chemical Society·Zahidul IslamAmnon Kohen
Apr 29, 2015·Physical Chemistry Chemical Physics : PCCP·Ananda K GhoshAmnon Kohen
Sep 17, 2008·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Ning ZhangWei-Shan Chen
Apr 28, 2019·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Cecilia PozziStefano Mangani
Jun 30, 2009·Comparative Biochemistry and Physiology. Toxicology & Pharmacology : CBP·Aldo A Arvizu-FloresRogerio R Sotelo-Mundo

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