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Biochemistry
11 05, 2024;63 (21): 2855-2867.

Noncovalent Inhibition and Covalent Inactivation of Proline Dehydrogenase by Analogs of -Propargylglycine.

John J Tanner, Juan Ji, Alexandra N Bogner, Gary K Scott, Sagar M Patel, Javier Seravalli, Kent S Gates, Christopher C Benz, Donald F Becker
Author Information
  1. John J Tanner: Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, United States. ORCID
  2. Juan Ji: Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, United States.
  3. Alexandra N Bogner: Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, United States. ORCID
  4. Gary K Scott: Buck Institute for Research on Aging, Novato, California 94945, United States.
  5. Sagar M Patel: Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, Nebraska 68588, United States. ORCID
  6. Javier Seravalli: Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, Nebraska 68588, United States.
  7. Kent S Gates: Department of Chemistry, University of Missouri, Columbia, Missouri 65211, United States. ORCID
  8. Christopher C Benz: Buck Institute for Research on Aging, Novato, California 94945, United States.
  9. Donald F Becker: Department of Biochemistry and Redox Biology Center, University of Nebraska-Lincoln, Lincoln, Nebraska 68588, United States. ORCID
PMID: 39437336 DOI: 10.1021/acs.biochem.4c00429

Abstract

The flavoenzyme proline dehydrogenase (PRODH) catalyzes the first step of proline catabolism, the oxidation of l-proline to Δ-pyrroline-5-carboxylate. The enzyme is a target for chemical probe discovery because of its role in the metabolism of certain cancer cells. -propargylglycine is the first and best characterized mechanism-based covalent inactivator of PRODH. This compound consists of a recognition module (glycine) that directs the inactivator to the active site and an alkyne warhead that reacts with the FAD after oxidative activation, leading to covalent modification of the FAD N5 atom. Here we report structural and kinetic data on analogs of -propargylglycine with the goals of understanding the initial docking step of the inactivation mechanism and to test the allyl group as a warhead. The crystal structures of PRODH complexed with unreactive analogs in which N is replaced by S show how the recognition module mimics the substrate proline by forming ion pairs with conserved arginine and lysine residues. Further, the C atom adjacent to the alkyne warhead is optimally positioned for hydride transfer to the FAD, providing the structural basis for the first bond-breaking step of the inactivation mechanism. The structures also suggest new strategies for designing improved -propargylglycine analogs. -allylglycine, which consists of a glycine recognition module and allyl warhead, is shown to be a covalent inactivator; however, it is less efficient than -propargylglycine in both enzyme inactivation and cellular assays. Crystal structures of the -allylglycine-inactivated enzyme are consistent with covalent modification of the N5 by propanal.

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Grants

  1. P30 GM124165/NIGMS NIH HHS
  2. R01 GM132640/NIGMS NIH HHS
  3. S10 OD021527/NIH HHS

MeSH Term

Glycine
Proline Oxidase
Kinetics
Crystallography, X-Ray
Humans
Catalytic Domain
Proline
Enzyme Inhibitors
Models, Molecular
Alkynes

Chemicals

Glycine
Proline Oxidase
propargylglycine
Proline
Enzyme Inhibitors
Alkynes
Journal Article Research Support, Non-U.S. Gov't Research Support, N.I.H., Extramural

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