This primary objective of the proposed research is to understand the control of the biochemical pathway for de novo biosynthesis of purine nucleotides. Specifically, this goal will be accomplished by determining the three-dimensional structure of glutamine PRPP amidotransferase, the enzyme that catalyzes the first and committed reaction in the pathway. Changes to the structure caused by inhibitor and activator molecules will also be determined. Most of the regulation of the purine biosynthetic pathway is directed towards this enzyme and a solid understanding of its structure and its reaction with effector molecules is critical to understanding regulation of the entire pathway. The health relatedness of the proposed research is twofold. Defects in the purine biosynthetic pathway have been associated with disorders such as hyperuricemia (gout), immunodeficiency and neurological symptoms. Secondly, components of the pathway, especially glutamine PRPP amidotransferase, are targets for the design of anticancer drugs due to the high rate of purine biosynthesis required by rapidly dividing cells. Glutamine PRPP amidotransferase contains an essential inorganic FeS center that apparently has a novel regulatory function and is not catalytic. The three-dimensional structure of the enzyme should clarify the role of this unusual metal center. The three-dimensional structure will be determined by X-ray crystallography and the structure determination should be a significant advance for the new crystallographic technology of multiwavelength anomalous dispersion. Successful application of this methodology to the glutamine PRPP amidotransferase structure will be a significant step towards proving its generality as a rapid and accurate means of structure determination for biological macromolecules.
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| Skiba, Meredith A; Maloney, Finn P; Dan, Qingyun et al. (2018) PKS-NRPS Enzymology and Structural Biology: Considerations in Protein Production. Methods Enzymol 604:45-88 |
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| Skiba, Meredith A; Sikkema, Andrew P; Moss, Nathan A et al. (2018) Biosynthesis of t-Butyl in Apratoxin A: Functional Analysis and Architecture of a PKS Loading Module. ACS Chem Biol 13:1640-1650 |
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| Skiba, Meredith A; Sikkema, Andrew P; Moss, Nathan A et al. (2017) A Mononuclear Iron-Dependent Methyltransferase Catalyzes Initial Steps in Assembly of the Apratoxin A Polyketide Starter Unit. ACS Chem Biol 12:3039-3048 |
| Maloney, Finn P; Gerwick, Lena; Gerwick, William H et al. (2016) Anatomy of the ?-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate. Proc Natl Acad Sci U S A 113:10316-21 |
| Stull, Frederick W; Bernard, Steffen M; Sapra, Aparna et al. (2016) Deprotonations in the Reaction of Flavin-Dependent Thymidylate Synthase. Biochemistry 55:3261-9 |
| Skiba, Meredith A; Sikkema, Andrew P; Fiers, William D et al. (2016) Domain Organization and Active Site Architecture of a Polyketide Synthase C-methyltransferase. ACS Chem Biol 11:3319-3327 |
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