New findings on SNP variants of human protein L-isoaspartyl methyltransferase that affect catalytic activity, thermal stability, and aggregation
| Autor: | Jean-Louis Bru, Baihe Chen, Eric Huynh, Dana W. Aswad, Jeungjin Kim, Mahsa Momen |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Aging Methyltransferase Molecular biology DNA Mutational Analysis Liquid Scintillation Counting Thermal Stability lcsh:Medicine Protein aggregation Biochemistry Database and Informatics Methods 0302 clinical medicine Gene Frequency Catalytic Domain Enzyme Stability Medicine and Health Sciences lcsh:Science Child chemistry.chemical_classification education.field_of_study Multidisciplinary Chemistry Physics Applied Mathematics Simulation and Modeling Temperature Melting Condensed Matter Physics Enzymes Bioassays and Physiological Analysis Knockout mouse Physical Sciences Thermodynamics Phase Transitions Algorithms Research Article Population DNA construction Research and Analysis Methods Polymorphism Single Nucleotide Protein Aggregation Pathological Catalysis 03 medical and health sciences Enzyme activator Protein Aggregates Protein D-Aspartate-L-Isoaspartate Methyltransferase Genetics SNP Humans education Allele frequency Aged Enzyme Assays Pharmacology lcsh:R Biology and Life Sciences Proteins Methyltransferases Pharmacologic-Based Diagnostics Enzyme Activation 030104 developmental biology Enzyme Genetics Population Biological Databases Molecular biology techniques Nerve Degeneration Mutation Databases Mutation Plasmid Construction Enzymology lcsh:Q Biochemical Analysis 030217 neurology & neurosurgery Mathematics |
| Zdroj: | PLoS ONE PLoS ONE, Vol 13, Iss 6, p e0198266 (2018) |
| ISSN: | 1932-6203 |
| Popis: | Protein L-isoaspartyl methyltransferase (PIMT/PCMT1), a product of the pcmt1 gene, catalyzes repair of abnormal L-isoaspartyl linkages in age-damaged proteins. Pcmt1 knockout mice exhibit a profound neuropathology and die 30-60 days postnatal from an epileptic seizure. Here we characterize four new SNP variants of human PIMT with respect to enzymatic activity, thermal stability, and propensity to aggregation. Under standard assay conditions, L191S, A150V, P174H and A65V showed activity losses of 72%, 64%, 61%, and 11% respectively. By differential scanning fluorimetry, melting temperature deviations were -5.2, -4.5, +0.5, and -3.4°C. SDS-PAGE of purified protein reveal significant aggregation of L191S, A150V, and P174H, but not A65V. We also report new data on three unusual PIMT variants among the 13 recently characterized by our laboratory. A7P and I58V were previously found to have 1.8-2.0 times the activity of WT PIMT in the standard assay; however, upon kinetic analysis, we find both variants exhibit reduced catalytic efficiency (Vmax/Km) due to weak isoaspartyl substrate binding. The near complete loss of activity ( |
| Databáze: | OpenAIRE |
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