ニワトリ4-ヒドロキシフェニルピルビン酸ジオキシゲナーゼ活性中心への4-ヒドロキシフェニルピルビン酸、α-ケトイソカプロン酸および2-(2-ニトロ-4-トリフルオロメチルベンゾイル)-シクロヘキサン-1,3ジオン(NTBC)の拮抗結合

概要

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We previously reported that the chicken 4-hydroxyphenylpyruvate dioxygenase (4-HPPD) also functions as a α-ketoisocaproate dioxygenase (α-KICD). To evaluate whether both substrates bind to the same active site, relation between the dual substrate property and active site (s) of chicken 4-HPPD was investigated using a recombinant chicken 4-HPPD. Co-presence of α-ketoisocaproate (α-KIC) resulted increasing of the Michaelis constant for 4-hydroxyphenylpyruvate (4-HPP) by four times without any significant change of the maximum velocity. 2-(2-nitro-4-trifluoromethylbenzoyl)-cyclohexane-1, 3-dione (NTBC), which is established to be a potent competitive inhibitor for mammalian 4-HPPD, inhibited competitively both 4-HPPD and α-KICD activities with the chicken expressed protein. Furthermore, NTBC concentration for 50% inhibition of 4-HPPD and α-KICD activities were 300nM for 4-HPP and 550nM for α-KIC, respectively. These results suggested strongly that both 4-HPP and α-KIC bind physiologically to the same active site of chicken 4-HPPD.