Functional analysis of human thromboxane synthase polymorphic variants
| Autor: | Richard J. Kulmacz, Elizabeth M. Poole, Chung Ying K. Chen, Cornelia M. Ulrich, Lee Ho Wang |
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| Rok vydání: | 2012 |
| Předmět: |
Models
Molecular Thromboxane Polymorphism Single Nucleotide Article Thromboxane receptor chemistry.chemical_compound Genetics Humans General Pharmacology Toxicology and Pharmaceutics Molecular Biology Genetics (clinical) biology Cytochrome P450 Malondialdehyde Molecular biology Thromboxane B2 Kinetics chemistry Biochemistry Biocatalysis biology.protein Prostaglandin H2 Molecular Medicine Electrophoresis Polyacrylamide Gel Mutant Proteins lipids (amino acids peptides and proteins) Thromboxane-A Synthase Cyclooxygenase Thromboxane-A synthase circulatory and respiratory physiology |
| Zdroj: | Pharmacogenetics and Genomics. 22:653-658 |
| ISSN: | 1744-6872 |
| DOI: | 10.1097/fpc.0b013e3283562d82 |
| Popis: | BACKGROUND Thromboxane A synthase (TXAS) metabolizes the cyclooxygenase product prostaglandin (PG) H2 into thromboxane H2 (TXA2), a potent inducer of blood vessel constriction and platelet aggregation. Nonsynonymous polymorphisms in the TXAS gene have the potential to alter TXAS activity and affect TXA2 generation. OBJECTIVES The aim of this study was to assess the functional effects of genetic variants in the TXAS protein, including K258E, L357V, Q417E, E450K, and T451N. METHODS Wild-type TXAS and the variant proteins were expressed in a bacterial system and purified by affinity and hydroxyapatite chromatography. The two characteristic catalytic activities of TXAS were assayed in each of the purified recombinant proteins: isomerization of PGH2 to TXA2 and fragmentation of PGH2 to 12-hydroxyheptadecatrienoic acid and malondialdehyde. RESULTS All of the variants showed both isomerization and fragmentation activities. The Km values of the variants ranged from 27 to 52 µmol/l PGH2 (wild-type value: 32 μmol/l PGH2); the Vmax values of the variants ranged from 18 to 40 U/mg (wild-type value: 41 U/mg). The kinetic differences were largest for the L357V variant, whose Vmax/Km ratio was just 27% of the wild-type value. CONCLUSION The increased Km and decreased Vmax values observed with L357V suggest that this variant may generate less TXA2 at the low levels of PGH2 expected in vivo, raising the possibility of attenuated signaling through the thromboxane pathway. |
| Databáze: | OpenAIRE |
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