In crystallo screening for proline analog inhibitors of the proline cycle enzyme PYCR1
Abstract
Pyrroline-5-carboxylate reductase 1 (PYCR1) catalyzes the biosynthetic half-reaction of the proline cycle by reducing Δ1-pyrroline-5-carboxylate (P5C) to proline through the oxidation of NAD(P)H. Many cancers alter their proline metabolism by up-regulating the proline cycle and proline biosynthesis, and knockdowns of PYCR1 lead to decreased cell proliferation. Thus, evidence is growing for PYCR1 as a potential cancer therapy target. Inhibitors of cancer targets are useful as chemical probes for studying cancer mechanisms and starting compounds for drug discovery; however, there is a notable lack of validated inhibitors for PYCR1. To fill this gap, we performed a small-scale focused screen of proline analogs using X-ray crystallography. Five inhibitors of human PYCR1 were discovered: l-tetrahydro-2-furoic acid, cyclopentanecarboxylate, l-thiazolidine-4-carboxylate, l-thiazolidine-2-carboxylate, and N-formyl l-proline (NFLP). The most potent inhibitor was NFLP, which had a competitive (with P5C) inhibition constant of 100 μm. The structure of PYCR1 complexed with NFLP shows that inhibitor binding is accompanied by conformational changes in the active site, including the translation of an α-helix by 1 Å. These changes are unique to NFLP and enable additional hydrogen bonds with the enzyme. NFLP was also shown to phenocopy the PYCR1 knockdown in MCF10A H-RASV12 breast cancer cells by inhibiting de novo prolinemore »
- Authors:
-
- Univ. of Missouri, Columbia, MO (United States)
- Katholieke Univ. Leuven, Heverlee (Belgium)
- Univ. of Nebraska, Lincoln, NE (United States)
- Publication Date:
- Research Org.:
- Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Org.:
- NIGMS; National Institutes of Health (NIH); USDOE Office of Science (SC)
- OSTI Identifier:
- 1765215
- Grant/Contract Number:
- AC02-05CH11231; AC02-06CH11357
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Journal of Biological Chemistry
- Additional Journal Information:
- Journal Volume: 295; Journal Issue: 52; Journal ID: ISSN 0021-9258
- Publisher:
- American Society for Biochemistry and Molecular Biology
- Country of Publication:
- United States
- Language:
- ENGLISH
- Subject:
- 60 APPLIED LIFE SCIENCES
Citation Formats
Christensen, Emily M., Bogner, Alexandra N., Vandekeere, Anke, Tam, Gabriela S., Patel, Sagar M., Becker, Donald F., Fendt, Sarah-Maria, and Tanner, John J. In crystallo screening for proline analog inhibitors of the proline cycle enzyme PYCR1. United States: N. p., 2020.
Web. doi:10.1074/jbc.ra120.016106.
Christensen, Emily M., Bogner, Alexandra N., Vandekeere, Anke, Tam, Gabriela S., Patel, Sagar M., Becker, Donald F., Fendt, Sarah-Maria, & Tanner, John J. In crystallo screening for proline analog inhibitors of the proline cycle enzyme PYCR1. United States. https://doi.org/10.1074/jbc.ra120.016106
Christensen, Emily M., Bogner, Alexandra N., Vandekeere, Anke, Tam, Gabriela S., Patel, Sagar M., Becker, Donald F., Fendt, Sarah-Maria, and Tanner, John J. Tue .
"In crystallo screening for proline analog inhibitors of the proline cycle enzyme PYCR1". United States. https://doi.org/10.1074/jbc.ra120.016106. https://www.osti.gov/servlets/purl/1765215.
@article{osti_1765215,
title = {In crystallo screening for proline analog inhibitors of the proline cycle enzyme PYCR1},
author = {Christensen, Emily M. and Bogner, Alexandra N. and Vandekeere, Anke and Tam, Gabriela S. and Patel, Sagar M. and Becker, Donald F. and Fendt, Sarah-Maria and Tanner, John J.},
abstractNote = {Pyrroline-5-carboxylate reductase 1 (PYCR1) catalyzes the biosynthetic half-reaction of the proline cycle by reducing Δ1-pyrroline-5-carboxylate (P5C) to proline through the oxidation of NAD(P)H. Many cancers alter their proline metabolism by up-regulating the proline cycle and proline biosynthesis, and knockdowns of PYCR1 lead to decreased cell proliferation. Thus, evidence is growing for PYCR1 as a potential cancer therapy target. Inhibitors of cancer targets are useful as chemical probes for studying cancer mechanisms and starting compounds for drug discovery; however, there is a notable lack of validated inhibitors for PYCR1. To fill this gap, we performed a small-scale focused screen of proline analogs using X-ray crystallography. Five inhibitors of human PYCR1 were discovered: l-tetrahydro-2-furoic acid, cyclopentanecarboxylate, l-thiazolidine-4-carboxylate, l-thiazolidine-2-carboxylate, and N-formyl l-proline (NFLP). The most potent inhibitor was NFLP, which had a competitive (with P5C) inhibition constant of 100 μm. The structure of PYCR1 complexed with NFLP shows that inhibitor binding is accompanied by conformational changes in the active site, including the translation of an α-helix by 1 Å. These changes are unique to NFLP and enable additional hydrogen bonds with the enzyme. NFLP was also shown to phenocopy the PYCR1 knockdown in MCF10A H-RASV12 breast cancer cells by inhibiting de novo proline biosynthesis and impairing spheroidal growth. In summary, we generated the first validated chemical probe of PYCR1 and demonstrated proof-of-concept for screening proline analogs to discover inhibitors of the proline cycle.},
doi = {10.1074/jbc.ra120.016106},
journal = {Journal of Biological Chemistry},
number = 52,
volume = 295,
place = {United States},
year = {Tue Dec 01 00:00:00 EST 2020},
month = {Tue Dec 01 00:00:00 EST 2020}
}
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