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Báo cáo khoa học: pH dependence, substrate specificity and inhibition of human kynurenine aminotransferase I

Human kynurenine aminotransferase I/glutamine trans-aminase K (hKAT-I) is an important multifunctional enzyme. This study systematically studies the substrates of hKAT-I and reassesses the effects of pH, Tris, amino acids anda-keto acids on the activity of the enzyme. The experi-ments were comprised of functional expression of the hKAT-I in an insect cell/baculovirus expression system, purification of its recombinant protein, and functional characterization of the purified enzyme. | Eur. J. Biochem. 271 4804-4814 2004 FEBS 2004 doi 10.1111 j.1432-1033.2004.04446.x pH dependence substrate specificity and inhibition of human kynurenine aminotransferase I Qian Han Junsuo Li and Jianyong Li Department of Pathobiology University of Illinois at Urbana-Champaign Urbana IL USA Human kynurenine aminotransferase I glutamine transaminase K hKAT-I is an important multifunctional enzyme. This study systematically studies the substrates of hKAT-I and reassesses the effects of pH Tris amino acids and a-keto acids on the activity of the enzyme. The experiments were comprised of functional expression of the hKAT-I in an insect cell baculovirus expression system purification of its recombinant protein and functional characterization of the purified enzyme. This study demonstrates that hKAT-I can catalyze kynurenine to kynurenic acid under physiological pH conditions indicates indo-3-pyruvate and cysteine as efficient inhibitors for hKAT-I and also provides biochemical information about the substrate specificity and cosubstrate inhibition of the enzyme. hKAT-I is inhibited by Tris under physiological pH conditions which explains why it has been concluded that the enzyme could not efficiently catalyze kynurenine transamination. Our findings provide a biochemical basis towards understanding the overall physiological role of hKAT-I in vivo and insight into controlling the levels of endogenous kynurenic acid through modulation of the enzyme in the human brain. Keywords cysteine indo-3-pyruvate kynurenic acid kynurenine aminotransferase pH effect. In mammals kynurenine aminotransferase I glutamine transaminase K EC 2.6.1.64 KAT-I is a multifunctional enzyme. In vitro the enzyme catalyzes the transamination of several amino acids e.g. glutamine methionine aromatic amino acids including kynurenine and also possesses cysteine S-conjugate b-lyase activity EC 4.4.1.13 1 . Kynurenic acid KYNA the stable product derived from the kynurenine transamination pathway 2-4 is .