pH dependence, substrate specificity and inhibition of human kynurenine aminotransferase I

Han, Qian; Li, Junsuo; Li, Jianyong

doi:10.1111/j.1432-1033.2004.04446.x

Cited by 97 publications

(145 citation statements)

References 43 publications

(78 reference statements)

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“…When the phosphate and borate buffer mixture, adjusted to pH 6.0 to 11.0, was used to prepare mKAT III-kynurenine-glyoxylate reaction mixtures, mKAT III displayed optimum activity around pH 9.0 to 10.0 (Fig. 1B), which is close to the optimum pH reported in the literature (3,4,16,49) for mammalian brain KAT I but apparently different from the optimum pH (7.5 to 9.0) of recombinant hKAT I (25). mKAT III was tested for aminotransferase activity toward 24 different amino acids, using glyoxylate as a primary amino group acceptor.…”

Section: Resultsmentioning

confidence: 52%

“…The substrate profile of mKAT III overlaps considerably with that of hKAT I (25), but hKAT I favors hydrophobic substrates and mKAT III prefers hydrophilic substrates. For example, mKAT III has high transamination activity toward asparagine, for which hKAT I has essentially no activity (25). Kinetic data provided an overview regarding the catalytic efficiency of mKAT III toward individual amino acids (Table 2).…”

Section: Resultsmentioning

confidence: 99%

“…In humans, rats, and mice, three proteins, designated kynurenine aminotransferases (KATs) I, II, and III, are involved in KYNA synthesis in the central nervous system (16,25,41,51,60). Very recently, mitochondrial aspartate aminotransferase from rat and human brains was reported to be able to catalyze the transamination of kynurenine to KYNA and therefore was named KAT IV (15).…”

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confidence: 99%

“…These studies suggest the importance of mammalian KAT I and KAT III in maintaining the KYNA level in the kat-2 Ϫ/Ϫ mouse brain. Although many studies have dealt with the biochemical characteristics of mammalian KAT I and KAT II (3,4,9,16,19,25,49), little is known about the biochemical activity of KAT III. Likewise, the crystal structures of human KAT I (hKAT I) (48) and its homologues, glutamine-phenylpyruvate aminotransferase from Thermus thermophilus HB8 (13) and KAT from a mosquito, Aedes aegypti (AeKAT) (22), have been determined, as well as the crystal structure of hKAT II (26,47) and its homologue from Pyrococcus horikoshii (7).…”

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confidence: 99%

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Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Han

Robinson

Cai

et al. 2009

Molecular and Cellular Biology

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Kynurenine aminotransferase III (KAT III) has been considered to be involved in the production of mammalian brain kynurenic acid (KYNA), which plays an important role in protecting neurons from overstimulation by excitatory neurotransmitters. The enzyme was identified based on its high sequence identity with mammalian KAT I, but its activity toward kynurenine and its structural characteristics have not been established. In this study, the biochemical and structural properties of mouse KAT III (mKAT III) were determined. Specifically, mKAT III cDNA was amplified from a mouse brain cDNA library, and its recombinant protein was expressed in an insect cell protein expression system. We established that mKAT III is able to efficiently catalyze the transamination of kynurenine to KYNA and has optimum activity at relatively basic conditions of around pH 9.0 and at relatively high temperatures of 50 to 60°C. In addition, mKAT III is active toward a number of other amino acids. Its activity toward kynurenine is significantly decreased in the presence of methionine, histidine, glutamine, leucine, cysteine, and 3-hydroxykynurenine. Through macromolecular crystallography, we determined the mKAT III crystal structure and its structures in complex with kynurenine and glutamine. Structural analysis revealed the overall architecture of mKAT III and its cofactor binding site and active center residues. This is the first report concerning the biochemical characteristics and crystal structures of KAT III enzymes and provides a basis toward understanding the overall physiological role of mammalian KAT III in vivo and insight into regulating the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

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Section: Resultsmentioning

confidence: 52%

Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Han

Robinson

Cai

et al. 2009

Molecular and Cellular Biology

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112

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“…However, they exhibit catalytic efficiencies quite different from those of the amino acids mentioned above. hKAT I is more efficient in transaminating hydrophobic amino acids (such as phenylalanine, leucine, and tryptophan), while AeKAT is more efficient in transaminating relatively hydrophilic amino acids (such as cysteine, histidine, tyrosine, and methionine) (4,5). Upon superposition of a dimer of the AeKATcysteine complex onto the hKAT I-phenylalanine structure ( Figure 6A), we identified the residues within 5 Å of both ligands and found that most residues were the same.…”

Section: Discussionmentioning

confidence: 99%

Structural Insight into the Mechanism of Substrate Specificity of Aedes Kynurenine Aminotransferase^,

et al. 2008

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Aedes aegypti kynurenine aminotransferase (AeKAT) is a multifunctional aminotransferase. It catalyzes the transamination of a number of amino acids and uses many biologically relevant α-keto acids as amino group acceptors. AeKAT also is a cysteine S-conjugate β-lyase. The most important function of AeKAT is the biosynthesis of kynurenic acid, a natural antagonist of NMDA and α7-nicotinic acetylcholine receptors. Here, we report the crystal structures of AeKAT in complex with its best amino acid substrates, glutamine and cysteine. Glutamine is found in both subunits of the biological dimer, and cysteine is found in one of the two subunits. Both substrates form external aldemines with pyridoxal 5-phosphate in the structures. This is the first instance in which one pyridoxal 5-phosphate enzyme has been crystallized with cysteine or glutamine forming external aldimine complexes, cysteinyl aldimine and glutaminyl aldimine. All the units with substrate are in the closed conformation form, and the unit without substrate is in the open form, which suggests that the binding of substrate induces the conformation change of AeKAT. By comparing the active site residues of the AeKAT-cysteine structure with those of the human KAT I-phenylalanine structure, we determined that Tyr286 in AeKAT is changed to Phe278 in human KAT I, which may explain why AeKAT transaminates hydrophilic amino acids more efficiently than human KAT I does.The sequence of Aedes aegypti kynurenine aminotransferase (AeKAT) 1 is 47% identical with that of human KAT I and 51.9% identical with that of human KAT III (1). Human, rat, and mouse KAT I enzymes have been extensively studied (2,3). Mammalian KAT I, also called glutamine transaminase K (GTK), is a multifunctional aminotransferase that acts on a number of amino acids (glutamine, cysteine, phenylalanine, kynurenine, methionine, etc.) and uses many biologically relevant keto acids as amino group acceptors (3,4). AeKAT and a glutamine:phenylpyruvate aminotransferase from Thermus thermophilus, homologues of mammalian KAT I, have also been systematically characterized (5,6). One of the enzymatic products of KAT I is kynurenic acid (KYNA), which is the only known endogenous antagonist of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors (7-9). KYNA is also the antagonist of the α7-nicotinic acetylcholine receptor (10-12). Very † This work was supported by Grant AI 44399 from the National Institutes of Health. ‡ The atomic coordinates and structure factors (entries 2r5c and 2r5e) have been deposited in the Protein Data Bank. Many other functions have been proposed for mammalian KAT I enzymes, including maintaining low levels of phenylpyruvate, closing the methionine salvage pathway, regulating α-keto acid levels, sparing the essential amino acids, and maintaining a continual equilibrium among the amino acids (3,17). However, quantitatively, the most important amine donor for KAT I enzymes in vivo is glutamine. The product of glutamine transamination (α-ketoglutaramate) is rapidly remov...

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Measurement of Cysteine S‐Conjugate β‐Lyase Activity

et al. 2010

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Cysteine S-conjugate β-lyases are pyridoxal 5′-phosphate (PLP)-containing enzymes that catalyze the conversion of cysteine S-conjugates [RSCH 2 CH(NH 3 + )CO 2 − ] and selenium Se-conjugates[RSeCH 2 CH(NH 3 + )CO 2 − ] that contain a leaving group in the β position to pyruvate, ammonium and a sulfur-containing fragment (RSH) or selenium-containing fragment (RSeH), respectively. At least ten PLP enzymes catalyze β-elimination reactions with such cysteine S-conjugates. All are enzymes involved in amino acid metabolism that do not normally catalyze a β-lyase reaction, but catalyze a non-physiological β-lyase side reaction that depends on the electron-withdrawing properties of the -SR or -SeR moiety. In the case of the cysteine S-conjugates, if the eliminated RSH is stable the compound may be S-thiomethylated and excreted (thiomethyl shunt) or Sglucuronidated and harmlessly excreted [the possibility that RSeH compounds may be similarly metabolized has not been extensively studied]. If, however, RSH is chemically reactive the cysteine S-conjugate may be toxic as a result of the β-lyase reaction. The cysteine S-conjugate β-lyase pathway is of particular interest to toxicologists because it is involved in the bioactivation (toxification) of halogenated alkenes and certain drugs. Keywords Ammonium; cysteine S-conjugates; cysteine S-conjugate β-lyases; S-(1,2-dichlorovinyl)-Lcysteine; S-(1,1,2,2-tetrafluoroethyl)-L-cysteine; pyruvateMany potentially toxic electrophiles, whether of endogenous origin (e.g. α,β-unsaturated aldehydes derived from the oxidation of lipids) or of exogenous origin (e.g. some drugs), are detoxified through the mercapturate pathway (Figure 1). In addition, some non-electrophilic toxins may be converted to electrophiles that are then detoxified through the mercapturate pathway. The electrophile is sequentially converted to the glutathione S-conjugate, cysteinylglycine S-conjugate, cysteinyl S-conjugate and N-acetyl cysteine S-conjugate (mercapturate) by the action of glutathione S-transferases, γ-glutamyltransferase (γ-glutamyltranspeptidase), dipeptidases and N-acetyl transferases, respectively. The mercapturate is generally more hydrophilic and water soluble than is the starting electrophile and is readily excreted in the urine or bile.If the cysteine S-conjugate contains a good leaving group (nucleofuge) in the β position the metabolism of the electrophile may be diverted irreversibly away from the mercapturate by the action of cysteine S-conjugate β-lyases. These PLP-containing enzymes catalyze the conversion of the cysteine S-conjugate [RSCH 2 CH(NH 3 + )CO 2 − ] to RSH and aminoacrylate

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pH dependence, substrate specificity and inhibition of human kynurenine aminotransferase I

Cited by 97 publications

References 43 publications

Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Structural Insight into the Mechanism of Substrate Specificity of Aedes Kynurenine Aminotransferase^,

Measurement of Cysteine S‐Conjugate β‐Lyase Activity

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pH dependence, substrate specificity and inhibition of human kynurenine aminotransferase I

Cited by 97 publications

References 43 publications

Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Biochemical and Structural Properties of Mouse Kynurenine Aminotransferase III

Structural Insight into the Mechanism of Substrate Specificity of Aedes Kynurenine Aminotransferase,

Measurement of Cysteine S‐Conjugate β‐Lyase Activity

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Structural Insight into the Mechanism of Substrate Specificity of Aedes Kynurenine Aminotransferase^,