Literature summary for 2.6.1.42 extracted from

Anderson, L.C.; Hakansson, M.; Walse, B.; Nilsson, C.L.
Intact protein analysis at 21 tesla and X-ray crystallography define structural differences in single amino acid variants of human mitochondrial branched-chain amino acid aminotransferase 2 (BCAT2) (2017), J. Am. Soc. Mass Spectrom., 28, 1796-1804 .

Application

Application	Comment	Organism
drug development	the enzyme mutant BCAT2 T186R is a precision drug target in glioblastoma (GBM)	Homo sapiens

Cloned(Commentary)

Cloned (Comment)	Organism
gene BCAT2, the BCAT2 protein is fused to an N-terminal His-tag for affinity purification, followed by a tobacco etch virus (TEV) protease cleavage site for tag removal, recombinant expression of His-tagged enzyme	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant detagged enzyme, mixing of 9.3 mg/ml protein in 25 mM HEPES, pH 7.5, 25 mM NaCl, 20 mM EDTA, 2.5% glycerol, and 20 mM DTT, is mixed with reservoir solution conataining 0.2 M NaF, 0.1 M Bis-Tris propane, pH 6.5, 20% w/v PEG 3350, and 10 mM DTT, a few days at 20°C, X-ray diffraction structure determination and analysis at 1.6 A, molecular replacement using the human BCAT2 BCAT2 inhibitor bound structure as a model (PDB ID 5CR5), model building	Homo sapiens

Crystallization (Comment)

Organism

purified recombinant detagged enzyme, mixing of 9.3 mg/ml protein in 25 mM HEPES, pH 7.5, 25 mM NaCl, 20 mM EDTA, 2.5% glycerol, and 20 mM DTT, is mixed with reservoir solution conataining 0.2 M NaF, 0.1 M Bis-Tris propane, pH 6.5, 20% w/v PEG 3350, and 10 mM DTT, a few days at 20°C, X-ray diffraction structure determination and analysis at 1.6 A, molecular replacement using the human BCAT2 BCAT2 inhibitor bound structure as a model (PDB ID 5CR5), model building

Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
T186R	site-directed mutagenesis, structure determination and comparison with wild-type enzyme	Homo sapiens

Localization

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

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	O15382	-	-

Purification (Commentary)

Purification (Comment)	Organism
recombinant N-terminally His-tagged BCAT2 by nickel affinity chromatography, tag cleavage by TEV protease, followed by gel filtration and ultrafiltration	Homo sapiens

Source Tissue

Source Tissue	Comment	Organism	Textmining
glioblastoma cell	-	Homo sapiens	-

Subunits

Subunits	Comment	Organism
?	x * 45000, SDS-PAGE	Homo sapiens
homodimer	2 * 41350, recombinant mutant T186R enzyme, mass spectrometry	Homo sapiens

Synonyms

Synonyms	Comment	Organism
Bcat2	-	Homo sapiens
branched-chain amino acid aminotransferase 2	-	Homo sapiens

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
20	-	assay at	Homo sapiens

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Homo sapiens

General Information

General Information	Comment	Organism
additional information	in silico structural homology modeling reveals that the Lys59 side-chain of BCAT2 may repulse the Arg186 in the variant protein (PDB ID 5MPR), leading to destabilization of the protein dimer and altered enzyme kinetics	Homo sapiens
physiological function	human mitochondrial branched-chain amino acid aminotransferase 2 is a putative factor of resistance of glioblastoma to standard-of-care-treatments. The enzyme generates glutamate, which is neurotoxic	Homo sapiens