5UAU

Structure of human PYCR-1 complexed with proline

Summary for 5UAU

• Entry DOI: 10.2210/pdb5uau/pdb
• Related: 5UAT 5UAV 5UAW 5UAX
• Descriptor: Pyrroline-5-carboxylate reductase 1, mitochondrial, PROLINE, SULFATE ION, ... (4 entities in total)
• Functional Keywords: amino-acid biosynthesis, oxidoreductase, proline biosynthesis
• Biological source: Homo sapiens (Human)
• Total number of polymer chains: 5
• Total formula weight: 171751.33

Authors

Tanner, J.J. (deposition date: 2016-12-20, release date: 2017-03-15, Last modification date: 2023-10-04)

Primary citation

Christensen, E.M.,Patel, S.M.,Korasick, D.A.,Campbell, A.C.,Krause, K.L.,Becker, D.F.,Tanner, J.J.
Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1.
J. Biol. Chem., 292:7233-7243, 2017

PubMed Abstract: 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.

PubMed: 28258219
DOI: 10.1074/jbc.M117.780288

Experimental method

X-RAY DIFFRACTION (1.9 Å)