title: Analysis of genetic polymorphism and biochemical characterization of a functionally decreased variant in prostacyclin synthase gene (CYP8A1) in humans.
abstract: Prostacyclin synthase (CYP8A1) is an enzyme responsible for the biosynthesis of prostacyclin (PGI2) which inhibits platelet activation and exhibits anti-inflammatory effect. The objectives of this study were to identify CYP8A1 genetic variants and characterize functional consequences of CYP8A1 variants. In total, 27 variants including four previously unidentified single-nucleotide polymorphisms (SNPs) were identified by direct DNA sequencing in Koreans (n=48). Among them, CYP8A1 A447T and E314Stop were newly assigned as CYP8A1(∗)5 and CYP8A1(∗)6 by the Human Cytochrome P450 Allele Nomenclature Committee, respectively. CYP8A1(∗)5 was found in the heme binding area in three individuals as a heterozygous mutation. To investigate the functional change of CYP8A1(∗)5, CYP8A1(∗)5 and wild-type CYP8A1 protein were overexpressed in an Escherichia coli expression system and purified. Metabolism of PGH2 by the CYP8A1(∗)5 protein exhibited significantly decreased activity, resulting in a 45% decrease in Vmax and a 1.8-fold decrease in intrinsic clearance compared to the wild-type. Based on the predicted crystal structure of CYP8A1(∗)5 using the Molecular Operating Environment platform, the distance from CYP8A1 Cys441 to the heme was altered with a significantly changed binding free energy for the mutant protein. Further studies would be needed to determine the effect of CYP8A1(∗)5 on PGI2 levels in humans.
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea.
authors.forename: Sun-Ah
authors.initials: SA
authors.lastname: Cho
authors.affiliation: Department of Pharmacology and Toxicology, University of Alabama at Birmingham, Birmingham, AL, USA.
authors.forename: Katie Jo
authors.initials: KJ
authors.lastname: Rohn-Glowacki
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea.
authors.forename: Yazun B
authors.initials: YB
authors.lastname: Jarrar
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea.
authors.forename: Myeongjin
authors.initials: M
authors.lastname: Yi
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea.
authors.forename: Woo-Young
authors.initials: WY
authors.lastname: Kim
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea; Department of Clinical Pharmacology, Inje University College of Medicine, Inje University Busan Paik Hospital, Busan 614-735, South Korea. Electronic address: phshinjg@gmail.com.
authors.forename: Jae-Gook
authors.initials: JG
authors.lastname: Shin
authors.affiliation: Department of Pharmacology, Pharmacogenomics Research Center, Inje University College of Medicine, Inje University, Busan 614-735, South Korea. Electronic address: 2sujun@inje.ac.kr.
authors.forename: Su-Jun
authors.initials: SJ
authors.lastname: Lee
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D018389:Codon, Nonsense; D003198:Computer Simulation; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004247:DNA / Q000235:genetics; D014644:Genetic Variation; D006418:Heme / Q000378:metabolism; D006579:Heterozygote; D006801:Humans; D007700:Kinetics; D015810:Linkage Disequilibrium; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D020641:Polymorphism, Single Nucleotide; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D056910:Republic of Korea; D017386:Sequence Homology, Amino Acid
chemical_list: D018389:Codon, Nonsense; D050505:Mutant Proteins; D011994:Recombinant Proteins; D006418:Heme; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; C472456:PTGIS protein, human
keywords: CYP8A1; Cytochrome P450; Polymorphism; Prostacyclin; SNP