Literature summary for 2.6.1.42 extracted from

Herbert, D.; Gibbs, S.; Riddick, A.; Conway, M.; Dong, M.
Crystal structure of an oxidized mutant of human mitochondrial branched-chain aminotransferase (2020), Acta Crystallogr. Sect. F, 76, 14-19 .

Cloned(Commentary)

Cloned (Comment)	Organism
recombinant expression of N-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3)	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant enzyme mutant C318A, hanging drop vapor diffusion method, mixing of 0.001 ml of 5 mg/ml protein in potassium phosphate, pH 7.5, with 0.001 ml of reservoir solution containing 200 mM magnesium acetate tetrahydrate, 100 mM sodium cacodylate trihydrate, pH 6.5, and 20% w/v PEG 8000, and equilibration against 0.5 ml of reservoir solution, 20°C, for oxidation of the C318A mutant, the protein crystals obtained are incubated in crystallization buffer with the addition of 1% hydrogen peroxide before applying the cryoprotectant, X-ray diffraction structure determination and analysis at 3.25 A resolution	Homo sapiens

Crystallization (Comment)

Organism

purified recombinant enzyme mutant C318A, hanging drop vapor diffusion method, mixing of 0.001 ml of 5 mg/ml protein in potassium phosphate, pH 7.5, with 0.001 ml of reservoir solution containing 200 mM magnesium acetate tetrahydrate, 100 mM sodium cacodylate trihydrate, pH 6.5, and 20% w/v PEG 8000, and equilibration against 0.5 ml of reservoir solution, 20°C, for oxidation of the C318A mutant, the protein crystals obtained are incubated in crystallization buffer with the addition of 1% hydrogen peroxide before applying the cryoprotectant, X-ray diffraction structure determination and analysis at 3.25 A resolution

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
C315A	site-directed mutagenesis, mutation of the redox sensor (Cys315) results in a significant loss of activity compared to wild-type	Homo sapiens
C318A	site-directed mutagenesis, the mutant shows unaltered activity compared to wild-type, structure analysis	Homo sapiens
C318A/C315CSD	the overall structure of the C318A/C315CSD variant overlays very well with that of the C318A mutant and the oxidized form of wild-type hBCATm, with a few exceptions in the interdomain loop (residues 171-181) and the N-terminal loop (residues 15-32)	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
mitochondrion	-	Homo sapiens	5739	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
L-leucine + 2-oxoglutarate	Homo sapiens	-	4-methyl-2-oxopentanoate + L-glutamate	-	r

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	O15382	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, anion exchange chromatography, and dialysis	Homo sapiens

Source Tissue

Source Tissue	Comment	Organism	Textmining
brain	-	Homo sapiens	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.	Reac.
L-leucine + 2-oxoglutarate	-	Homo sapiens	4-methyl-2-oxopentanoate + L-glutamate	-	r

Subunits

Subunits	Comment	Organism
homodimer	-	Homo sapiens

Synonyms

Synonyms	Comment	Organism
Bcat2	-	Homo sapiens
branched-chain aminotransferase	-	Homo sapiens
hBCAT	-	Homo sapiens
hBCATm	-	Homo sapiens

Cofactor

Cofactor	Comment	Organism	Structure
pyridoxal 5'-phosphate	PLP, dependent on, the reaction is accompanied by the interconversion of the cofactor pyridoxal 5'-phosphate (PLP) and pyridoxamine 5'-phosphate. PLP is linked to Lys202 by a Schiff base	Homo sapiens

General Information

General Information	Comment	Organism
malfunction	mutation of the redox sensor (Cys315) results in a significant loss of activity, with no loss of activity reported on the mutation of the resolving cysteine (Cys318), which allows the reversible formation of a disulfide bond between Cys315 and Cys318	Homo sapiens
additional information	active site structure	Homo sapiens
physiological function	human branched-chain aminotransferase (hBCAT) catalyzes the transamination of the branched-chain amino acids leucine, valine and isoleucine and 2-oxoglutarate to their respective 2-oxo acids and glutamate. hBCAT activity is regulated by a CXXC center located approx. 10 A from the active site. This redox-active center facilitates recycling between the reduced and oxidized states, representing hBCAT in its active and inactive forms, respectively. The structure reveals the modified CXXC center in a conformation similar to that in the oxidized wild type, supporting the notion that its regulatory mechanism depends on switching the Cys315 side chain between active and inactive conformations. The enzyme plays a significant role in amino-acid metabolism and whole-body nitrogen shuttling, in particular with respect to the de novo synthesis of the neurotransmitter glutamate in the brain	Homo sapiens