Functional analysis of three CYP1A2 variants found in a Japanese population

Yoshiro Saito, Nobumitsu Hanioka, Keiko Maekawa, Takashi Isobe, Yumi Tsuneto, Ryosuke Nakamura, Akiko Soyama, Shogo Ozawa, Toshiko Tanaka-Kagawa, Hideto Jinno, Shizuo Narimatsu, Jun Ichi Sawada

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Abstract

Human cytochrome P450 1A2 (CYP1A2) catalyzes the metabolism of many important drugs and environmental chemicals. We previously reported three naturally occurring genetic polymorphisms (125C>G, Pro42Arg, CYP1A2*15; 1130G>A, Arg377Gln, *16; and 1367G>A, Arg456His, *8) found in a Japanese population. In this study, these variant enzymes were expressed in Chinese hamster V79 cells, and their mRNA and protein expression levels as well as catalytic activities were determined. All three variant enzymes showed reduced protein expression levels (66% for Pro42Arg and approximately 30% for Arg377Gln and Arg456His) compared with that of the wild type (WT) without any change in mRNA expression levels. Kinetic analysis for 7-ethoxyresorufin O-deethylation revealed that Vmax and Vmax/Km of all three variants were less than 3 and 1% of the WT, respectively, although the Km value was significantly increased only in the Arg377Gln variant (approximately a 9-fold increase). Markedly reduced activities of the three variants were also observed for phenacetin O-deethylation. In the reduced CO difference spectral analysis using recombinant proteins produced in the Sf21/baculovirus system, the peak at 450 nm seen in the WT protein was hardly observed in the three variants, suggesting marked reductions in their hemoprotein formation. These results suggest that Pro42, Arg377, and Arg456 are critical amino acids for the production of catalytically active CYP1A2 holoenzyme.

Original languageEnglish
Pages (from-to)1905-1910
Number of pages6
JournalDrug Metabolism and Disposition
Volume33
Issue number12
DOIs
Publication statusPublished - Dec 2005

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Cytochrome P-450 CYP1A2
Functional analysis
Phenacetin
Population
Messenger RNA
Holoenzymes
Proteins
Baculoviridae
Genetic Polymorphisms
Enzymes
Carbon Monoxide
Cricetulus
Polymorphism
Recombinant Proteins
Metabolism
Spectrum analysis
Catalyst activity
Amino Acids
Kinetics
Pharmaceutical Preparations

ASJC Scopus subject areas

  • Pharmacology
  • Toxicology

Cite this

Saito, Y., Hanioka, N., Maekawa, K., Isobe, T., Tsuneto, Y., Nakamura, R., ... Sawada, J. I. (2005). Functional analysis of three CYP1A2 variants found in a Japanese population. Drug Metabolism and Disposition, 33(12), 1905-1910. https://doi.org/10.1124/dmd.105.005819

Functional analysis of three CYP1A2 variants found in a Japanese population. / Saito, Yoshiro; Hanioka, Nobumitsu; Maekawa, Keiko; Isobe, Takashi; Tsuneto, Yumi; Nakamura, Ryosuke; Soyama, Akiko; Ozawa, Shogo; Tanaka-Kagawa, Toshiko; Jinno, Hideto; Narimatsu, Shizuo; Sawada, Jun Ichi.

In: Drug Metabolism and Disposition, Vol. 33, No. 12, 12.2005, p. 1905-1910.

Research output: Contribution to journalArticle

Saito, Y, Hanioka, N, Maekawa, K, Isobe, T, Tsuneto, Y, Nakamura, R, Soyama, A, Ozawa, S, Tanaka-Kagawa, T, Jinno, H, Narimatsu, S & Sawada, JI 2005, 'Functional analysis of three CYP1A2 variants found in a Japanese population', Drug Metabolism and Disposition, vol. 33, no. 12, pp. 1905-1910. https://doi.org/10.1124/dmd.105.005819
Saito Y, Hanioka N, Maekawa K, Isobe T, Tsuneto Y, Nakamura R et al. Functional analysis of three CYP1A2 variants found in a Japanese population. Drug Metabolism and Disposition. 2005 Dec;33(12):1905-1910. https://doi.org/10.1124/dmd.105.005819
Saito, Yoshiro ; Hanioka, Nobumitsu ; Maekawa, Keiko ; Isobe, Takashi ; Tsuneto, Yumi ; Nakamura, Ryosuke ; Soyama, Akiko ; Ozawa, Shogo ; Tanaka-Kagawa, Toshiko ; Jinno, Hideto ; Narimatsu, Shizuo ; Sawada, Jun Ichi. / Functional analysis of three CYP1A2 variants found in a Japanese population. In: Drug Metabolism and Disposition. 2005 ; Vol. 33, No. 12. pp. 1905-1910.
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abstract = "Human cytochrome P450 1A2 (CYP1A2) catalyzes the metabolism of many important drugs and environmental chemicals. We previously reported three naturally occurring genetic polymorphisms (125C>G, Pro42Arg, CYP1A2*15; 1130G>A, Arg377Gln, *16; and 1367G>A, Arg456His, *8) found in a Japanese population. In this study, these variant enzymes were expressed in Chinese hamster V79 cells, and their mRNA and protein expression levels as well as catalytic activities were determined. All three variant enzymes showed reduced protein expression levels (66{\%} for Pro42Arg and approximately 30{\%} for Arg377Gln and Arg456His) compared with that of the wild type (WT) without any change in mRNA expression levels. Kinetic analysis for 7-ethoxyresorufin O-deethylation revealed that Vmax and Vmax/Km of all three variants were less than 3 and 1{\%} of the WT, respectively, although the Km value was significantly increased only in the Arg377Gln variant (approximately a 9-fold increase). Markedly reduced activities of the three variants were also observed for phenacetin O-deethylation. In the reduced CO difference spectral analysis using recombinant proteins produced in the Sf21/baculovirus system, the peak at 450 nm seen in the WT protein was hardly observed in the three variants, suggesting marked reductions in their hemoprotein formation. These results suggest that Pro42, Arg377, and Arg456 are critical amino acids for the production of catalytically active CYP1A2 holoenzyme.",
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