. This program investigates the role of protein'protein interactions in carrier protein-dependent biosynthesis, including fatty acid synthase, polyketide synthase, and non-ribosomal peptide synthetase pathways. The natural products associated with these pathways serve as therapeutics including antibiotics, antifungals, and anticancer agents. The ability to direct the biosynthesis of these pathways for the production of untried bioactive compounds is of critical importance for new and improved therapies. In prior years, we demonstrated how protein'protein interactions between carrier proteins and partner protein domains direct reactivity and guide processivity. Moreover, we developed a suite of chemical biology tools to stabilize and interrogate these interactions at atomic resolution through innovative crosslinker development and structural biology. We now propose to expand these tools with new, caged crosslinkers designed to capture transient partner proteins. With this comprehensive library of crosslinkers, we will evaluate carrier protein interactions with partner proteins that include ketoreductases, enoyl reductases, and thioesterases from fatty acid and polyketide synthases, ketosynthase/chain-length factors and acyltransferases from polyketide synthases, and condensation domains, halogenases, and oxidases from non-ribosomal peptide synthetases. Once captured, these crosslinked species will be studied by solution-phase NMR, kinetic and thermodynamic assays, X-ray crystallography, and molecular dynamics simulations to more fully elucidate mechanism and specificity conferred by carrier protein-substrate/intermediate/produ interactions. Finally, we will use data collected in this program as an informatic platform to enable synthetic biological production of new natural product hybrids.

Public Health Relevance

. Natural products provide critical medicines to treat human disease; therefore, an understanding of their biosynthesis will directly support development of new and improved natural product-based therapeutics. Developing on outstanding progress in recent years towards revealing the fundamental protein?protein interactions between biosynthetic enzymes, this proposal aims to further our knowledge through the development of tools to structurally characterize biosynthetic machinery at atomic resolution with temporal acuity. These studies will provide a foundation to rationally manipulate these pathways, affording access to new molecular species for next-generation drug discovery and development.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Macromolecular Structure and Function A Study Section (MSFA)
Program Officer
Bond, Michelle Rueffer
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University of California, San Diego
Schools of Arts and Sciences
La Jolla
United States
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Sarria, Stephen; Bartholow, Thomas G; Verga, Adam et al. (2018) Matching Protein Interfaces for Improved Medium-Chain Fatty Acid Production. ACS Synth Biol 7:1179-1187
Chen, Aochiu; Re, Rebecca N; Burkart, Michael D (2018) Type II fatty acid and polyketide synthases: deciphering protein-protein and protein-substrate interactions. Nat Prod Rep 35:1029-1045
Dick, Benjamin L; Patel, Ashay; McCammon, J Andrew et al. (2017) Effect of donor atom identity on metal-binding pharmacophore coordination. J Biol Inorg Chem 22:605-613
Finzel, Kara; Beld, Joris; Burkart, Michael D et al. (2017) Utilizing Mechanistic Cross-Linking Technology to Study Protein-Protein Interactions: An Experiment Designed for an Undergraduate Biochemistry Lab. J Chem Educ 94:375-379
Jaremko, Matt J; Lee, D John; Patel, Ashay et al. (2017) Manipulating Protein-Protein Interactions in Nonribosomal Peptide Synthetase Type II Peptidyl Carrier Proteins. Biochemistry 56:5269-5273
Barajas, Jesus F; Shakya, Gaurav; Moreno, Gabriel et al. (2017) Polyketide mimetics yield structural and mechanistic insights into product template domain function in nonreducing polyketide synthases. Proc Natl Acad Sci U S A 114:E4142-E4148
Vickery, Christopher R; La Clair, James J; Burkart, Michael D et al. (2016) Harvesting the biosynthetic machineries that cultivate a variety of indispensable plant natural products. Curr Opin Chem Biol 31:66-73
Mindrebo, Jeffrey T; Nartey, Charisse M; Seto, Yoshiya et al. (2016) Unveiling the functional diversity of the alpha/beta hydrolase superfamily in the plant kingdom. Curr Opin Struct Biol 41:233-246
McCulloch, Ian P; La Clair, James J; Jaremko, Matt J et al. (2016) Fluorescent Mechanism-Based Probe for Aerobic Flavin-Dependent Enzyme Activity. Chembiochem 17:1598-601
Rivera Jr, Heriberto; Dhar, Sachin; La Clair, James J et al. (2016) An unusual intramolecular trans-amidation. Tetrahedron 72:3605-3608

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