Identification and functional characterization of novel CYP2J2 variants: G312R variant causes loss of enzyme catalytic activity

Abstract

CYP2J2 plays important roles in the metabolism of therapeutic drugs, such as astemizole and ebastine, as well as endogenous fatty acids. This study aimed to identify CYP2J2 genetic variants in Koreans and to characterize their functional consequences. From direct sequencing of the CYP2J2 gene, 12 genetic variations, including the two novel nonsynonymous mutations G312R and P351L, were identified from 93 Korean subjects. The two novel CYP2J2 variants were co-expressed with NADPH-cytochrome P450 reductase in Sf9 cells and their catalytic activities were quantified. The recombinant CYP2J2 G312R variant showed almost complete loss of enzymatic activity, as determined by CYP2J2-catalysed astemizole O-demethylation and ebastine hydroxylation. The CYP2J2 P351L variant showed enzymatic activities that were comparable with the wild-type CYP2J2. The reduced CO spectra of the recombinant CYP2J2 proteins suggested no CO binding to the heme in CYP2J2 G312R. In addition, molecular modelling of the three-dimensional structure consistently predicted that there might be spatial hindrance between heme and the bulky side chain of the R312 residue in CYP2J2 G312R variant. The CYP2J2 G312R variant was not found in 192 Chinese, 99 African-Americans, 100 Caucasians and 159 Vietnamese subjects. Two of the 192 Chinese subjects (0.52%) were heterozygous for CYP2J2 P351L. Twelve CYP2J2 variants, including two novel nonsynonymous variants, were identified in a Korean population. The G312R variant is the first nonfunctional CYP2J2 allele to be identified, and is expected to influence the disposition of its substrate therapeutics, as well as endogenous compounds.

Identification and Functional Assessment ofBCRPPolymorphisms in a Korean Population

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TheCYP3A4*18Allele, the Most Frequent Coding Variant in Asian Populations, Does Not Significantly Affect the Midazolam Disposition in Heterozygous Individuals

“…CYP3A4 content was determined by difference spectroscopy, as described previously (Omura and Sato, 1964). The expression levels of the CYP3A4.1, CYP3A4.18, and CPR proteins were verified by Western blot analysis, as described previously (Lee et al, 2005b).…”

Screening and identification of peritoneal metastasis-related genes of gastric adenocarcinoma using a cDNA microarray

“…When gene chip was applied in the beginning, the quality of microarray was not satisfactory from clone preparation, chip production, variations from hybridization, or inadequate data analysis. However, with improved chip quality and hybridization technique, recent microarray data showed good concordance with RT-PCR or Northern blot method with reliable biological meanings (Lee et al, 2005;Davidson et al, 2006). In the present study, upregulation of the S100A4 and CTNNB1 genes or downregulation of the PTEN gene in GC9811-P were also observed using RT-PCR analysis.…”