Structural Basis for the Substrate Inhibition of Proline Utilization A by Proline

Korasick, David; Pemberton, Travis; Arentson, Benjamin; Becker, Donald; Tanner, John

doi:10.3390/molecules23010032

Structural Basis for the Substrate Inhibition of Proline Utilization A by Proline

Journal Article · Sat Dec 23 00:00:00 EST 2017 · Molecules

DOI:https://doi.org/10.3390/molecules23010032· OSTI ID:1628485

Korasick, David ^[1]; Pemberton, Travis ^[2]; Arentson, Benjamin ^[3]; Becker, Donald ^[3]; Tanner, John ^[2]

Univ. of Missouri, Columbia, MO (United States); DOE/OSTI
Univ. of Missouri, Columbia, MO (United States)
Univ. of Nebraska, Lincoln, NE (United States)

Proline utilization A (PutA) is a bifunctional flavoenzyme that catalyzes the two-step oxidation of l-proline to l-glutamate using spatially separated proline dehydrogenase (PRODH) and l-glutamate-γ-semialdehyde dehydrogenase (GSALDH) active sites. Substrate inhibition of the coupled PRODH-GSALDH reaction by proline is a common kinetic feature of PutAs, yet the structural basis for this phenomenon remains unknown. To understand the mechanism of substrate inhibition, we determined the 2.15 Å resolution crystal structure of Bradyrhizobium japonicum PutA complexed with proline. Proline was discovered in five locations remote from the PRODH active site. Most notably, strong electron density indicated that proline bound tightly to the GSAL binding site of the GSALDH active site. The pose and interactions of proline bound in this site are remarkably similar to those of the natural aldehyde substrate, GSAL, implying that proline inhibits the GSALDH reaction of PutA. Kinetic measurements show that proline is a competitive inhibitor of the PutA GSALDH reaction. Together, the structural and kinetic data show that substrate inhibition of the PutA coupled reaction is due to proline binding in the GSAL site.

View Accepted Manuscript (DOE)

Research Organization:: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: National Institutes of Health (NIH), NIGMS; USDOE Office of Science (SC)

Grant/Contract Number:: AC02-05CH11231

OSTI ID:: 1628485

Journal Information:: Molecules, Journal Name: Molecules Journal Issue: 1 Vol. 23; ISSN MOLEFW; ISSN 1420-3049

Publisher:: MDPICopyright Statement

Country of Publication:: United States

Language:: English

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37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
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substrate inhibition
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Structural Basis for the Substrate Inhibition of Proline Utilization A by Proline

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