title: Effect of R119G Mutation on Human P5CR1 Dynamic Property and Enzymatic Activity.
abstract: Pyrroline-5-carboxylate reductase (P5CR1) is a universal housekeeping enzyme that catalyzes the reduction of Δ1-pyrroline-5-carboxylate (P5C) to proline with concomitant oxidation of NAD(P)H to NAD(P)+. The enzymatic cycle between P5C and proline is important for function in amino acid metabolism, apoptosis, and intracellular redox potential balance in mitochondria. Autosomal recessive cutis laxa (ARCL) results from a mutation in P5CR1 encoded by PYCR1. Specifically, the R119G mutation is reported to be linked to ARCL although it has not yet been characterized. We synthesized R119G P5CR1 and compared it to WT P5CR1. Foldx prediction of WT and R119G mutant P5CR1 protein stability suggests that the R119G mutation could significantly reduce protein stability. We also performed enzymatic activity assays to determine how the mutation impacts P5CR1 enzymatic function. The results of these experiments show that mutagenesis of R119 to G decreases P5CR1 catalytic efficiency for 3,4-dehydro-L-proline relative to WT. Mutagenesis and kinetic studies reveal that the activity of the mutant decreases as temperature increases from 5°C to 37°C, with almost no activity at 37°C, indicating that this mutation impairs P5CR1 function in vivo. Conversely, WT P5CR1 retains its activity after incubation at 37°C and has essentially no remaining activity at 75°C. Taken together, our experimental results indicate the R119G mutation could be an involving pathomechanism for ARCL.
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Linhua
authors.initials: L
authors.lastname: Li
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Yujia
authors.initials: Y
authors.lastname: Ye
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Sang
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Yirui
authors.initials: Y
authors.lastname: Yin
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Hu
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Deyun
authors.initials: D
authors.lastname: Li
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Wen
authors.initials: W
authors.lastname: Wan
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Rui
authors.initials: R
authors.lastname: Li
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Longjun
authors.initials: L
authors.lastname: Li
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Linling
authors.initials: L
authors.lastname: Ma
authors.affiliation: Department of Computer Science, The Faculty of Basic Medicine, Kunming Medical University, Kunming, China.
authors.forename: Yuehui
authors.initials: Y
authors.lastname: Xie
authors.affiliation: Laboratory of Molecular Cardiology, Department of Cardiology, The First Affiliated Hospital of Kunming Medical University, Kunming, China.
authors.forename: Zhaohui
authors.identifier: 0000-0003-0490-8748
authors.initials: Z
authors.lastname: Meng
mesh_terms: D019943:Amino Acid Substitution; D002384:Catalysis; D018360:Crystallography, X-Ray; D003483:Cutis Laxa / Q000201:enzymology / Q000235:genetics; D004795:Enzyme Stability / Q000235:genetics; D006801:Humans; D020125:Mutation, Missense; D017510:Protein Folding; D011762:Pyrroline Carboxylate Reductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000097502:delta-1-Pyrroline-5-Carboxylate Reductase
chemical_list: D011762:Pyrroline Carboxylate Reductases; D000097502:delta-1-Pyrroline-5-Carboxylate Reductase