Genetic Polymorphisms of the HumanCYP2A13Gene: Identification of Single-Nucleotide Polymorphisms and Functional Characterization of an Arg257Cys Variant

Abstract

Human cytochrome P450 2A13 (CYP2A13), which is highly efficient in the metabolic activation of a major tobacco-specific carcinogen, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), may play important roles in xenobiotic toxicity and tobacco-related tumorigenesis in the respiratory tract. The aim of this study was to identify any genetic polymorphisms of the CYP2A13 gene, which may alter the metabolic capacities of the enzyme. Polymerase chain reaction (PCR) single-strand conformational polymorphism analysis was used to identify single-nucleotide polymorphisms (SNPs) in all of the exons and at the exon-intron boundaries, and PCR-restriction fragment length polymorphism analysis and DNA sequencing were used to determine the frequencies of the newly identified variant alleles in the four major ethnic groups. Blood spot DNA from more than 100 individuals was used for these analyses. Seven variant alleles were found, but only one SNP was detected in the coding region, in exon 5, leading to an Arg257Cys amino acid change. The frequencies of the Arg257Cys allele in white, black, Hispanic, and Asian individuals are 1.9%, 14.4%, 5.8%, and 7.7%, respectively. Functional analysis of the variant protein was performed following its heterologous expression. The Arg257Cys variant was 37 to 56% less active than the wild-type Arg-257 protein toward all substrates tested. With NNK, Cys-257 had higher K m and lower V max values than did Arg-257, with a Ͼ2-fold decrease in catalytic efficiency. The Arg257Cys mutation could provide some protection against xenobiotic toxicity in the respiratory tract to individuals who are homozygous for the Cys-257 allele.

Catalog of 680 variations among eight cytochrome P450 (CYP) genes, nine esterase genes, and two other genes in the Japanese population

“…Among the CYP genes examined here, other investigators have previously detected 27 polymorphisms that would affect amino acid sequences [ten in CYP2A6, six in CYP2B6, three in CYP2E, and eight in TBXAS1; Human Cytochrome P450 (CYP) Allele Nomenclature Committee, http://www.imm.ki.se/CYPalleles/]. Zhang et al (2002) have detected an additional SNP (Arg257-Cys) in the coding region of CYP2A13. However, of the 28 polymorphisms reported previously, we have found only six in our Japanese population sample (Ile471Thr in CYP2A6; Arg257Cys in CYP2A13; Arg22Cys, Gln172His, and Arg487Cys in CYP2B6; Glu450Lys in TBXAS1).…”

“…Open boxes Exons, hatching unsequenced regions of repetitive elements, ATG initiation codon, TGA, TAG stop codons Ariyoshi et al (2001) c SNP previously reported by Zhang et al (2002) d SNP previously reported by Lang et al (2001) e SNPs previously reported by Fairbrother et al (1998) f SNP previously reported by Baek et al (1996) g SNP previously reported by Chevalier et al (2001) Gene and variation no. …”

“…Zhang et al (2002) have identified a C-to-T polymorphism (Arg257Cys) in exon 5 of the gene, and the product of this variant is 37% to 56% less active than the wild-type protein toward all substrates tested.…”

Genetic susceptibility of lung cancer associated with common variants in the 3′ untranslated regions of the adenosine triphosphate‐binding cassette B1 (ABCB1) and ABCC1 candidate transporter genes for carcinogen export

“…For example, we and other groups have documented that the interaction of CYP2A13 and NNK is associated with lung cancer. 4,10,11 In addition, we previously analyzed the possible association between lung cancer and variants of the b-2 adrenergic receptor gene ADRB2, which is the key mediator of the NNK nongenotoxic pathway. 12 It also has been reported that polymorphisms in the phase II conjugation enzyme genes, such as uridine diphosphate glucuronosyltransferase 1 family, polypeptide A7 (UGT1A7) and UGT2B17, have implications in lung cancer etiology.…”

The Effects of Single Nucleotide Polymorphisms in CYP2A13 on Metabolism of 5-Methoxypsoralen

“…These substitutions can affect substrate specificity or metabolic turnover of the CYP2A13 enzyme. However, functional differences in the SNP variants of CYP2A13 are limited; the studies using recombinant proteins expressed in mammalian cell cultures showed that the CYP2A13*4 protein has decreased metabolic activity on NNK and aflatoxin B1, and that the CYP2A13*2 protein is less efficient in metabolic activation of NNK than wild-type CYP2A13*1 (Zhang et al, 2002;Wang et al, 2003Wang et al, , 2006.…”