Literature summary for 2.6.1.42 extracted from

Conway, M.E.
Emerging moonlighting functions of the branched-chain aminotransferase proteins (2020), Antioxid. Redox Signal., 34, 1048-1067.

Cloned(Commentary)

Cloned (Comment)	Organism
gene BCAT1, located at 12p12.1, the BCAT1 gene is a target for c-myc	Homo sapiens
gene BCAT2, located at 19q13.33	Homo sapiens

Crystallization (Commentary)

Crystallization (Comment)	Organism
crystal structure analyses, PDB IDs 1KTA, 1KT8, and 2HG8	Homo sapiens

Protein Variants

Protein Variants	Comment	Organism
C315A	analysis of the PLP-bound mutant enzyme crystal structure, ODB ID 2HG8	Homo sapiens
C335S/C338S	dimerization of BCATc occurs concurrently with GSNO-mediated S-glutathionylation, which is abolished when substituted with the double mutant C335S/C338S	Homo sapiens

Inhibitors

Inhibitors	Comment	Organism
benzothiazolyl-L-cysteine	BTC, is a substrate and inactivator of BCATm but not BCATc, but BTC inhibits transamination between leucine and 2-oxoglutarate for both enzymes; BTC, is a substrate and inactivator of BCATm but not BCATc, but BTC inhibits transamination between leucine and 2-oxoglutarate for both enzymes. Inactivation of BCATm can occur through the interaction of aminoacrylate generated from the beta-lyase reaction with the PLP cofactor, through alkylation of a key residue at the active site, or through thioacylation by the eliminated sulphur-containing fragment	Homo sapiens
beta-chloro-L-alanine	the compound is able to fit into the active site of BCATm, leading to inhibition of BCATm	Homo sapiens
trichloroethylene/dichloroacetylene	DCVC, selectively inhibits BCATc, resulting in suppressed transamination and subsequent neurotoxicity	Homo sapiens

Localization

Localization	Comment	Organism	GeneOntology No.	Textmining
cytosol	-	Homo sapiens	5829	-
mitochondrion	-	Homo sapiens	5739	-

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
L-isoleucine + 2-oxoglutarate	Homo sapiens	-	3-methyl-2-oxopentanoate + L-glutamate	-	r
L-leucine + 2-oxoglutarate	Homo sapiens	-	4-methyl-2-oxopentanoate + L-glutamate	-	r
L-valine + 2-oxoglutarate	Homo sapiens	-	3-methyl-2-oxobutanoate + L-glutamate	-	r

Organism

Organism	UniProt	Comment	Textmining
Homo sapiens	O15382	-	-
Homo sapiens	P54687	-	-

Oxidation Stability

Oxidation Stability	Organism
BCATm is more susceptible to ROS, whereas modification of BCATc by RNS results in S-nitrosylation and S-glutathionylation	Homo sapiens

Posttranslational Modification

Posttranslational Modification	Comment	Organism
additional information	the enzyme is subject to posttranslational modification by reactive oxygen and nitrogen species	Homo sapiens
S-nitrosylation	of the BCAT proteins at the CXXC redox-active motif	Homo sapiens
side-chain modification	S-glutathionylation of the BCAT proteins at the CXXC redox-active motif	Homo sapiens
side-chain modification	S-glutathionylation of the BCAT proteins at the CXXC redox-active motif. Dimerization of BCATc occurs concurrently with GSNO-mediated S-glutathionylation, which is abolished when substituted with the double mutant C335S/C338S. S-glutathionylation of BCATc increases as the environment became more oxidizing. S-glutathionylation of BCATc is a mechanism to preserve BCAT integrity under cellular stress	Homo sapiens

Reaction

Reaction	Comment	Organism	Reaction ID
L-leucine + 2-oxoglutarate = 4-methyl-2-oxopentanoate + L-glutamate	transamination functions through a ping pong kinetic mechanism involving two half-reactions, characterised by the interconversion of its PLP and PMP (pyridoxamine) form. In the first half reaction, binding of the BCAA triggers transaldimination releasing the active lysine and an external aldimine is formed between substrate and the PLP cofactor. The lysine residue is the catalytic base in the next step, a 1,3-prototropic shift that converts the external aldimine into a ketamine intermediate. Here, a proton is abstracted from the ?-carbon of the substrate amino acid and subsequent re-protonation of the aldehyde carbon of the coenzyme yields the ketamine. The final step in the first half-reaction is the hydrolysis of the ketamine to give the PMP form of the enzyme and an 2-oxo acid. The second half-reaction is the reversal of the first half-reaction with a different 2-oxo acid. The committed step in the complete oxidation of the BCAA is catalysed by the BCKD generating acyl CoAs	Homo sapiens	a
L-leucine + 2-oxoglutarate = 4-methyl-2-oxopentanoate + L-glutamate	transamination functions through a ping pong kinetic mechanism involving two half-reactions, characterised by the interconversion of its PLP and PMP (pyridoxamine) form. In the first half reaction, binding of the BCAA triggers transaldimination releasing the active lysine and an external aldimine is formed between substrate and the PLP cofactor. The lysine residue is the catalytic base in the next step, a 1,3-prototropic shift that converts the external aldimine into a ketamine intermediate. Here, a proton is abstracted from the alpha-carbon of the substrate amino acid and subsequent re-protonation of the aldehyde carbon of the coenzyme yields the ketamine. The final step in the first half-reaction is the hydrolysis of the ketamine to give the PMP form of the enzyme and an 2-oxo acid. The second half-reaction is the reversal of the first half-reaction with a different 2-oxo acid. The committed step in the complete oxidation of the BCAA is catalysed by the BCKD generating acyl CoAs. During transamination, the BCAA substrate binds to the PLP cofactor at the bottom of the active site and donates its nitrogen to PLP-BCATm	Homo sapiens	a

Source Tissue

Source Tissue	Comment	Organism	Textmining
brain	-	Homo sapiens	-
brain	in human brain, BCATc is solely expressed in glutamatergic and GABAergic neurons in all brain regions examined. In the hippocampal and temporal region of the brain, intense staining of BCATc is reported in the neuronal cell bodies indicating that the role of BCATc is to contribute to the glutamate pool rather than excitation. A role in neurotransmitter release during excitation is also proposed as BCATc is localised along the axons. Isozyme BCAT1 reguation in the brain, overview	Homo sapiens	-
breast cancer cell	BCATc and BCATm are increased in breast cancer	Homo sapiens	-
carcinoma cell	BCAT1 has been extensively reported in malignancies including gliomas, ovarian, colorectal, gastric cancer, nasoparyngeal carcinomas, breast cancer, and chronic myeloid leukemia (CML)	Homo sapiens	-
glioma cell	-	Homo sapiens	-
hippocampus	-	Homo sapiens	-
kidney	-	Homo sapiens	-
liver	-	Homo sapiens	-
lymphocyte	-	Homo sapiens	-
macrophage	-	Homo sapiens	-
additional information	highest expression of BCATm reported is in the pancreas followed by muscle, brain, and kidney in descending order. Skeletal muscle is a major site for BCAA metabolism, facilitated by BCATm	Homo sapiens	-
additional information	the cytosolic form (BCATc) is largely brain specific with expression reported also in the peripheral nervous system, placenta, testis and ovaries. Activated lymphocytes and macrophages also show expression of BCATc. BCAT1 is highly expressed early in embryogenesis and in c-myc-based tumours. Although BCATc is highly expressed in tumours, normally it shows cell specific expression particular to glutamatergic and GABAergic neurons	Homo sapiens	-
muscle	-	Homo sapiens	-
neuron	neuronal cell body	Homo sapiens	-
ovary	-	Homo sapiens	-
pancreas	-	Homo sapiens	-
peripheral nervous system	-	Homo sapiens	-
placenta	-	Homo sapiens	-
skeletal muscle	-	Homo sapiens	-
testis	-	Homo sapiens	-

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
L-isoleucine + 2-oxoglutarate	-	Homo sapiens	3-methyl-2-oxopentanoate + L-glutamate	-	r
L-leucine + 2-oxoglutarate	-	Homo sapiens	4-methyl-2-oxopentanoate + L-glutamate	-	r
L-valine + 2-oxoglutarate	-	Homo sapiens	3-methyl-2-oxobutanoate + L-glutamate	-	r
additional information	in addition to this traditional role, BCAT possesses other activities including its role as a cysteine S-conjugate beta-lyase and a thiol disulfide isomerase. Aminotransferase proteins including BCAT have been shown to catalyse beta-lyase reactions with amino acids containing a good leaving group in the beta position. Possible BCAT oxidoreductase activity	Homo sapiens	?	-	-
additional information	in addition to this traditional role, BCAT possesses other activities including its role as a cysteine S-conjugate beta-lyase and a thiol disulfide isomerase. Aminotransferase proteins including BCAT have been shown to catalyse beta-lyase reactions with amino acids containing a good leaving group in the beta position. Possible BCAT oxidoreductase activity. BCAT enhances the overall PDI activity and increases the rate of refolding. BCATm may be a chaperone for PDI operating through a thiol disulfide exchange mechanism or independently as an oxidoreductase regulated through the redox environment	Homo sapiens	?	-	-

Subunits

Subunits	Comment	Organism
homodimer	2 * 41730, sequence calculation	Homo sapiens
homodimer	2* 42800, about, sequence calculation	Homo sapiens
More	BCATm has an interdomain loop. Each monomer of BCATm is composed of a small (residues 1-175) and a large domain (residues 176-365) with an interdomain loop. At the dimer interface two PLP cofactors reside	Homo sapiens
More	the BCATc polypeptide consists of a small (residues 1-188) and large domain (residues 202-C-terminus), connected by an interdomain loop (residues 189-201). Three-dimensional enzyme structure complexed with gabapentin in the oxidised and reduced state	Homo sapiens

Synonyms

Synonyms	Comment	Organism
BcaT	-	Homo sapiens
BCAT1	-	Homo sapiens
Bcat2	-	Homo sapiens
BCATc	-	Homo sapiens
BCATm	-	Homo sapiens
branched-chain aminotransferase	-	Homo sapiens

Cofactor

Cofactor	Comment	Organism
additional information	analysis of enzyme bound structures of pyridoxal 5'-phosphate and pyridoxamine phosphate, overview	Homo sapiens
pyridoxal 5'-phosphate	PLP, the PLP cofactor is bound to the active site Lys207 of BCAT by an internal aldimine, with a Schiff base	Homo sapiens
pyridoxal 5'-phosphate	PLP, the PLP cofactor is located at the bottom of the active site with its re-face toward the protein side, forming a covalent Schiff base with the side chain of the catalytic Lys222	Homo sapiens

General Information

General Information	Comment	Organism
malfunction	with respect to BCATc, single point mutations of all six detectable thiols show that those in the CXXC motif, C335 and C338, have the most impact on BCAT activity. Steady state kinetics show that mutation of the thiol at position C335 had the largest effect relative to all other single point mutations. For all amino acid substrates there is a significant decrease in kcat/Km values. The other non-CXXC cysteine mutants show differential effects on turnover, with the most significant observed for C221S. When the thiol at position C338 is substituted an increase in peroxide sensitivity is observed, indicating that C335 is the redox sensor	Homo sapiens
metabolism	the enzyme is part of the branched-chain amino acids metabolism, redox regulation of BCAT, detailed overview. Redox regulation of BCATm is important for substrate channelling. For association to occur, BCATm needs to be in its reduced-PLP form and both proteins in their open structure. In the PLP-form, binding of BCATm to E1 increases the kinetic rate of decarboxylation of the BCKAs, whereas no binding occurs when BCATm is in the PMP form	Homo sapiens
metabolism	the enzyme is part of the branched-chain amino acids metabolism, redox regulation of BCAT, residue C335 is the redox sensor, detailed overview. Like whole body transamination, neurotransmitter synthesis can be finely regulated through dietary BCAAs. S-glutathionylation of BCATc is a mechanism to preserve BCAT integrity under cellular stress	Homo sapiens
additional information	the redox active CXXC motif is unique to the branched-chain aminotransferase (BCAT) proteins, the CXXC motif is at the heart of the catalytic center and functions through thiol disulfide exchange. Importance of the CXXC motif in regulating BCAT activity under hypoxic conditions, a characteristic of tumours. The key active-site residues involved in substrate and PLP binding are identical between isoforms, with the exception of one residue (Val336 in BCATc is Gln in BCATm), which does not explain the catalytic or regulatory differences presented for the BCAT proteins. BCATs contain a redox active CXXC center, analysis of the functional role	Homo sapiens
additional information	the redox active CXXC motif is unique to the branched-chain aminotransferase (BCAT) proteins. The CXXC motif is at the heart of the catalytic center and functions through thiol disulfide exchange. Importance of the CXXC motif in regulating BCAT activity under hypoxic conditions, a characteristic of tumours. The key active-site residues involved in substrate and PLP binding are identical between isoforms, with the exception of one residue (Val336 in BCATc is Gln in BCATm), which does not explain the catalytic or regulatory differences presented for the BCAT proteins. BCATs contain a redox active CXXC center, analysis of the functional role	Homo sapiens
physiological function	the BCAT proteins have been assigned an additional thiol oxidoreductase activity that can accelerate the refolding of proteins, in particular when S-glutathionylated, supporting a chaperone role for BCAT in protein folding. Interplay of the redox regulation of BCAT with important cell signalling mechanisms. The two isozymes BCAT1 and 2 have similar substrate specificities, but their regulation, tissue specific expression and compartmentation together with their response to different redox environments point to alternate functions for these proteins. The mitochondrial form is responsible for the majority of transamination in the body due to its ubiquitous expression in almost every tissue excluding the liver. High levels of BCAAs may exacerbate the generation of sorbitol accumulation. Moreover, increased levels of BCAAs have also been shown to reduce fatty acid oxidation in muscles, leading to the accumulation of acylcarnitines and insulin resistance. The metabolic imbalances include a role of BCATm and its regulation through changes in the redox environment. BCAT enhances the overall PDI activity and increases the rate of refolding. BCATm may be a chaperone for PDI operating through a thiol disulfide exchange mechanism or independently as an oxidoreductase regulated through the redox environment. BCATm may operate in a neuroprotective capacity, as an auxiliary mechanism to the endothelial system to support glutamate efflux. The redox state of BCAT signals differential phosphorylation by protein kinase C regulating the trafficking of cellular pools of BCAT	Homo sapiens
physiological function	the BCAT proteins have been assigned an additional thiol oxidoreductase activity that can accelerate the refolding of proteins, in particular when S-glutathionylated, supporting a chaperone role for BCAT in protein folding. Interplay of the redox regulation of BCAT with important cell signalling mechanisms. The two isozymes BCAT1 and 2 have similar substrate specificity, but their regulation, tissue specific expression and compartmentation together with their response to different redox environments point to alternate functions for these proteins. In human brain, BCATc is solely expressed in glutamatergic and GABAergic neurons in all brain regions examined. In the hippocampal and temporal region of the brain, intense staining of BCATc is reported in the neuronal cell bodies indicating that the role of BCATc is to contribute to the glutamate pool rather than excitation. A role in neurotransmitter release during excitation is also proposed as BCATc is localised along the axons. Roles of isozyme BCAT1 in cancer, detailed overview	Homo sapiens