Response to boguslaw stec's letter to the editor of <i>protein science</i>

Celeste, L.R.; Lovelace, L.L.; Lebioda, Lukasz

doi:10.1002/pro.2251

Cited by 2 publications

(3 citation statements)

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“…Many examples of incorrectly built structures or misplaced ligands have appeared in the crystallographic literature over the past few years (33)(34)(35)(36). Some have arisen from updates in genomic databases that revealed older structures solved with incorrect sequences (33,37,38). Others are due to the very large number of new structures appearing each year coupled with the fact that many of them come from laboratories with little crystallographic experience (39).…”

Section: Enzymementioning

confidence: 99%

Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1

Christensen¹,

Patel

Korasick

et al. 2017

Journal of Biological Chemistry

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Pyrroline-5-carboxylate reductase (PYCR) is the final enzyme in proline biosynthesis, catalyzing the NAD(P)H-dependent reduction of Δ-pyrroline-5-carboxylate (P5C) to proline. Mutations in the gene alter mitochondrial function and cause the connective tissue disorder cutis laxa. Furthermore, PYCR1 is overexpressed in multiple cancers, and the knock-out suppresses tumorigenic growth, suggesting that PYCR1 is a potential cancer target. However, inhibitor development has been stymied by limited mechanistic details for the enzyme, particularly in light of a previous crystallographic study that placed the cofactor-binding site in the C-terminal domain rather than the anticipated Rossmann fold of the N-terminal domain. To fill this gap, we report crystallographic, sedimentation-velocity, and kinetics data for human PYCR1. Structures of binary complexes of PYCR1 with NADPH or proline determined at 1.9 Å resolution provide insight into cofactor and substrate recognition. We see NADPH bound to the Rossmann fold, over 25 Å from the previously proposed site. The 1.85 Å resolution structure of a ternary complex containing NADPH and a P5C/proline analog provides a model of the Michaelis complex formed during hydride transfer. Sedimentation velocity shows that PYCR1 forms a concentration-dependent decamer in solution, consistent with the pentamer-of-dimers assembly seen crystallographically. Kinetic and mutational analysis confirmed several features seen in the crystal structure, including the importance of a hydrogen bond between Thr-238 and the substrate as well as limited cofactor discrimination.

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Section: Enzymementioning

confidence: 99%

Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1

Christensen¹,

Patel

Korasick

et al. 2017

Journal of Biological Chemistry

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“…However, inspection of the deposited data (https://doi.org/10.2210/pdb4IOJ/pdb, https://doi.org/10.2210/pdb4IOK/pdb, https://doi.org/10.2210/pdb4IOL/pdb and https://doi.org/10.2210/pdb4IOM/pdb) revealed deficiencies, and additional questions about the validity of these structures had been raised [56]. New refinements [57] were carried out and now the refined models (https://doi.org/10.2210/pdb4JIM/pdb, https://doi.org/10.2210/pdb4JJK/pdb, https://doi.org/10.2210/pdb4JJZ/pdb and https://doi.org/10.2210/pdb4JKI/pdb) provide us with templates to explore the reaction mechanism of the human MTHFD1 synthetase domain.…”

Section: Mthfd1mentioning

confidence: 99%

“…However, inspection of the deposited data (4IOJ, 4IOK, 4IOL and 4IOM) revealed deficiencies, and additional questions about the validity of these structures had been raised [56]. New refinements [57] were carried out and now the refined models (4JIM, 4JJK, 4JJZ and 4JKI) provide us with templates to explore the reaction mechanism of the human MTHFD1 synthetase domain.…”

Section: Mthfd1mentioning

confidence: 99%

Pairing structural reconstruction with catalytic competence to evaluate the mechanisms of key enzymes in the folate‐mediated one‐carbon pathway

Zhao

Kaldis

2022

The FEBS Journal

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Mammalian metabolism comprises a series of interlinking pathways that include two major cycles: the folate and methionine cycles. The folate‐mediated metabolic cycle uses several oxidation states of tetrahydrofolate to carry activated one‐carbon units to be readily used and interconverted within the cell. They are required for nucleotide synthesis, methylation and metabolism, and particularly for proliferation of cancer cells. Based on the latest progress in genome‐wide CRISPR loss‐of‐function viability screening of 789 cell lines, we focus on the most cancer‐dependent enzymes in this pathway, especially those that are hyperactivated in cancer, to provide new insight into the chemical basis for cancer drug development. Since the complete 3D structure of several of these enzymes of the one‐carbon pathway in their active form are not available, we used homology modelling integrated with the interpretation of the reaction mechanism. In addition, have reconstructed the most likely scenario for the reactions taking place paired with their catalytic competence that provides a testable framework for this pathway.

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Response to boguslaw stec's letter to the editor of protein science

Abstract: PDB Code(s): http://firstglance.jmol.org/fg.htm?mol=4JIM, http://firstglance.jmol.org/fg.htm?mol=4JJZ, http://firstglance.jmol.org/fg.htm?mol=4JJK, http://firstglance.jmol.org/fg.htm?mol=4JKI

Cited by 2 publications

References 0 publications

Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1

Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1

Pairing structural reconstruction with catalytic competence to evaluate the mechanisms of key enzymes in the folate‐mediated one‐carbon pathway

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