Effect of cytochrome P450 2C19 and 2C9 amino acid residues 72 and 241 on metabolism of tricyclic antidepressant drugs.
Although cytochromes P450 2C9 (CYP2C9) and 2C19 (CYP2C19) have 91% amino acid identity, they have different substrate specificities. Previous studies have suggested that several amino acid residues may be involved in substrate specificity. In this study, we focused on the roles of two amino acids, residues 72 and 241. The amino acids in these positions have opposite charges in CYP2C9 and 2C19; the former has lysines in both positions (Lys72 and Lys241), and the latter has glutamic acids (Glu72 and Glu241). Reciprocal mutants for both CYP2C19 and 2C9 were produced, and their metabolic activities and spectroscopic properties were examined using three tricyclic antidepressant (TCA) drugs: amitriptyline, imipramine, and dothiepin. Although CYP2C19 wild-type (WT) had a high metabolic activity for all three drugs, the E72K mutation decreased enzymatic activity by 29-37%, while binding affinities were diminished 2.5- to 20-fold. On the other hand, low activity and low affinity of CYP2C9 WT were recovered notably by K72E mutation. The metabolic activities and binding affinities were minimally affected by CYP2C19 E241K and CYP2C9 K241E mutations. We could also show linear correlations between metabolic activities and binding affinities, and hence we conclude that amino acid residue 72 plays a key role in TCA drug metabolism by limiting the binding affinities of CYP2C19 and CYP2C9.
Graduate School of Pharmaceutical Sciences, Osaka University.
Tamer Zekry
TZ
Attia
Taku
T
Yamashita
Mohamed Abdelkhalek
MA
Hammad
Akinori
A
Hayasaki
Takumi
T
Sato
Masayoshi
M
Miyamoto
Yuki
Y
Yasuhara
Takashi
T
Nakamura
Yusuke
Y
Kagawa
Hirofumi
H
Tsujino
Mahmoud Ahmed
MA
Omar
Osama Hassan
OH
Abdelmageed
Sayed Mohamed
SM
Derayea
Tadayuki
T
Uno
D000595:Amino Acid Sequence; D000639:Amitriptyline / Q000378:metabolism*; D000929:Antidepressive Agents, Tricyclic / Q000378:metabolism*; D001189:Aryl Hydrocarbon Hydroxylases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D065731:Cytochrome P-450 CYP2C19; D065729:Cytochrome P-450 CYP2C9; D004308:Dothiepin / Q000378:metabolism*; D006801:Humans; D007099:Imipramine / Q000378:metabolism*; D008969:Molecular Sequence Data; D017354:Point Mutation; D011485:Protein Binding; D013379:Substrate Specificity
D000929:Antidepressive Agents, Tricyclic; D000639:Amitriptyline; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; D065731:Cytochrome P-450 CYP2C19; D007099:Imipramine; D004308:Dothiepin