Dual substrate recognition of aminotransferases

Hirotsu, Ken; Goto, Masaru; Okamoto, Atsushi; Miyahara, Ikuko

doi:10.1002/tcr.20042

Cited by 110 publications

(100 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This substrate-mediated conformation change is also observed in other aminotransferases (21,33,41). The AeKATcysteine and AeKAT-glutamine structures show that the protein subunits are crystallized as closed forms when they bind substrates.…”

Section: Discussionmentioning

confidence: 73%

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

et al. 2008

View full text Add to dashboard Cite

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...

show abstract

Section: Discussionmentioning

confidence: 73%

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

et al. 2008

View full text Add to dashboard Cite

show abstract

“…A possible explanation would be the substrate recognition mechanism of the transaminase subgroup II enzymes. In general, the substrate in the active site is recognized by the two substrate binding pockets around the PLPlysine Schiff base (9,23). The main roles of the binding pockets are to recognize the side chain of the substrate and to anchor the carboxylate groups of the substrate (9, 39).…”

Section: Resultsmentioning

confidence: 99%

Cloning and Characterization of a Novel β-Transaminase fromMesorhizobiumsp. Strain LUK: a New Biocatalyst for the Synthesis of Enantiomerically Pure β-Amino Acids

Kim

Kyung

Yun

et al. 2007

Appl Environ Microbiol

View full text Add to dashboard Cite

A novel ␤-transaminase gene was cloned from Mesorhizobium sp. strain LUK. By using N-terminal sequence and an internal protein sequence, a digoxigenin-labeled probe was made for nonradioactive hybridization, and a 2.5-kb gene fragment was obtained by colony hybridization of a cosmid library. Through Southern blotting and sequence analysis of the selected cosmid clone, the structural gene of the enzyme (1,335 bp) was identified, which encodes a protein of 47,244 Da with a theoretical pI of 6.2. The deduced amino acid sequence of the ␤-transaminase showed the highest sequence similarity with glutamate-1-semialdehyde aminomutase of transaminase subgroup II. The ␤-transaminase showed higher activities toward D-␤-aminocarboxylic acids such as 3-aminobutyric acid, 3-amino-5-methylhexanoic acid, and 3-amino-3-phenylpropionic acid. The ␤-transaminase has an unusually broad specificity for amino acceptors such as pyruvate and ␣-ketoglutarate/ oxaloacetate. The enantioselectivity of the enzyme suggested that the recognition mode of ␤-aminocarboxylic acids in the active site is reversed relative to that of ␣-amino acids. After comparison of its primary structure with transaminase subgroup II enzymes, it was proposed that R43 interacts with the carboxylate group of the ␤-aminocarboxylic acids and the carboxylate group on the side chain of dicarboxylic ␣-keto acids such as ␣-ketoglutarate and oxaloacetate. R404 is another conserved residue, which interacts with the ␣-carboxylate group of the ␣-amino acids and ␣-keto acids. The ␤-transaminase was used for the asymmetric synthesis of enantiomerically pure ␤-aminocarboxylic acids. (3S)-Amino-3-phenylpropionic acid was produced from the ketocarboxylic acid ester substrate by coupled reaction with a lipase using 3-aminobutyric acid as amino donor.

show abstract

“…(i) Principle of the method Alanine Transaminase Activity Assay Kit (colorimetric) is a rapid and simple assay where ALT catalysis the transfer of an amino group from alanine to α-ketoglutarate, the products of this reversible transamination reaction being pyruvate and glutamate [12,13]. The kit provides a rapid, simple, sensitive, and reliable test suitable for high throughput activity assay of ALT with a detection limit of 10mU per well [14].…”

Section: Methods Used For Ast Determinationmentioning

confidence: 99%

The Investigation of Zinc-Dependent Enzymes in Pregnant Women and their Correlation to Zinc Deficiency

Treska¹

2015

IJSQA

View full text Add to dashboard Cite

Abstract:Zinc is present throughout the body in low concentration, but in most tissues. It performs multiple critical functions and must be supplied at adequate levels consistently or deficiency states will result, from mild to severe. Zinc deficiency is insufficient zinc to meet the needs of biological organisms. Due to its essentiality, a lack of this trace element leads to far more severe and widespread problems. Enzymes are large biological molecules responsible for the thousands of chemical inter-conversions that sustain life. Enzymes are usually very specific as to which reactions they catalyze and the substrates that are involved in these reactions. Complementary shape, charge and hydrophilic/ hydrophobic characteristics of enzymes and substrates are responsible for this specificity. Specimens were collected at the University Hospital of Obstetrics and Gynecology "Queen Geraldine" in Tirana, Albania during a period of time from year 2011 to 2013. We took into consideration 500 cases of pregnant women from first to third trimester of pregnancy. Serum zinc was measured by using Photometry (End-Point) method, whereas zinc enzymes such as Alkaline Phosphatase, Amylase, Gamma-glutamyl Transpeptidase, Lactate Dehydrogenase, Creatine Kinase, Aspartate Amino-Transferase, Glutamic Pyruvic Transaminase, Lipase measured with the corresponding methods: Beckman Synchron LX20, Colorimetric ab102523 kit, GenWays GGT kit, nonradioactive colorimetric LDH kit, Max Discovery, Colorimetric kit, Abcam kit and Biovision kit. From 500 cases taken into consideration, 190 pregnant women (38%) had normal zinc values (70 -120 mcg/dl) 58 pregnant women (11.6%) had zinc levels between 60 -69.9 mcg/dl, patients which were identified as pregnant women with zinc deficiency, as a consequence of oral contraceptive use; 252 pregnant women (50.4%) with zinc values < 60 mcg/dl, identified as patients with zinc deficiency, a result of malnutrition and also urinary elimination of digestive liquids; 56 pregnant women (11.2%) with zinc values < 30 mcg/dl, identified as patient with definite deficiency as a result of different diseases like acrodermatitis enteropathica ect. According to these results, there was a positive correlation between zinc and enzymes such as ALP, CK and LDH in pregnant women suffering from hypertension, whereas a negative correlation of zinc to enzymes such GGT, AST and GPT in cases with anemia. Zinc prophylactic treatment is important before and during pregnancy. Without a proper nutritional requirement the person falls in the state of zinc deficiency.

show abstract

Dual substrate recognition of aminotransferases

Cited by 110 publications

References 36 publications

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

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

Cloning and Characterization of a Novel β-Transaminase fromMesorhizobiumsp. Strain LUK: a New Biocatalyst for the Synthesis of Enantiomerically Pure β-Amino Acids

The Investigation of Zinc-Dependent Enzymes in Pregnant Women and their Correlation to Zinc Deficiency

Contact Info

Product

Resources

About

Dual substrate recognition of aminotransferases

Cited by 110 publications

References 36 publications

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

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

Cloning and Characterization of a Novel β-Transaminase fromMesorhizobiumsp. Strain LUK: a New Biocatalyst for the Synthesis of Enantiomerically Pure β-Amino Acids

The Investigation of Zinc-Dependent Enzymes in Pregnant Women and their Correlation to Zinc Deficiency

Contact Info

Product

Resources

About

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

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