Human mitochondrial and cytosolic branched-chain aminotransferases are cysteine S-conjugate beta-lyases, but turnover leads to inactivation.
The mitochondrial and cytosolic branched-chain aminotransferases (BCAT(m) and BCAT(c)) are homodimers in the fold type IV class of pyridoxal 5'-phosphate-containing enzymes that also contains D-amino acid aminotransferase and 4-amino-4-deoxychorismate lyase (a beta-lyase). Recombinant human BCAT(m) and BCAT(c) were shown to have beta-lyase activity toward three toxic cysteine S-conjugates [S-(1,1,2,2-tetrafluoroethyl)-L-cysteine, S-(1,2-dichlorovinyl)-L-cysteine, and S-(2-chloro-1,1,2-trifluoroethyl)-L-cysteine] and toward beta-chloro-L-alanine. Human BCAT(m) is a much more effective beta-chloro-L-alanine beta-lyase than two aminotransferases (cytosolic and mitochondrial isozymes of aspartate aminotransferase) previously shown to possess this activity. BCAT(m), but not BCAT(c), also exhibits measurable beta-lyase activity toward a relatively bulky cysteine S-conjugate [benzothiazolyl-L-cysteine]. Benzothiazolyl-L-cysteine, however, inhibits the L-leucine-alpha-ketoglutarate transamination reaction catalyzed by both enzymes. Inhibition was more pronounced with BCAT(m). In the presence of beta-lyase substrates and alpha-ketoisocaproate (the alpha-keto acid analogue of leucine), no transamination could be detected. Therefore, with an amino acid containing a good leaving group in the beta position, beta-elimination is greatly preferred over transamination. Both BCAT isozymes are rapidly inactivated by the beta-lyase substrates. The ratio of turnover to inactivation per monomer in the presence of toxic halogenated cysteine S-conjugates is approximately 170-280 for BCAT(m) and approximately 40-50 for BCAT(c). Mitochondrial enzymes of energy metabolism are especially vulnerable to thioacylation and inactivation by the reactive fragment released from toxic, halogenated cysteine S-conjugates such as S-(1,1,2,2-tetrafluoroethyl)-L-cysteine. The present results suggest that BCAT isozymes may contribute to the mitochondrial toxicity of these compounds by providing thioacylating fragments, but inactivation of the BCAT isozymes might also block essential metabolic pathways.
Cooper
Arthur J L
AJ
Department of Biochemistry, Weill Medical College of Cornell University, New York, NY 10021, USA. acooper@burke.org
Bruschi
Sam A
SA
Conway
Myra
M
Hutson
Susan M
SM
D000641:Ammonia / Q000378:metabolism; D013437:Carbon-Sulfur Lyases / Q000378:metabolism*; D003545:Cysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D003600:Cytosol / Q000201:enzymology; D006219:Halogens / Q000737:chemistry; D006801:Humans; D006845:Hydrocarbons, Fluorinated / Q000494:pharmacology; D007651:Keto Acids / Q000494:pharmacology; D007930:Leucine / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D019289:Pyruvic Acid / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature; D000637:Transaminases / Q000378:metabolism*; D015091:beta-Alanine / Q000031:analogs & derivatives* / Q000494:pharmacology; D005466:Fluorocarbons
D000641:Ammonia; D013437:Carbon-Sulfur Lyases; D003545:Cysteine; D006219:Halogens; D006845:Hydrocarbons, Fluorinated; D007651:Keto Acids; D007930:Leucine; D019289:Pyruvic Acid; D011994:Recombinant Proteins; D000637:Transaminases; D015091:beta-Alanine; C006977:3-chloroalanine; C013082:alpha-ketoisocaproic acid; C064116:S-(1,1,2,2-tetrafluoroethyl)cysteine; D005466:Fluorocarbons; C019798:branched-chain-amino-acid transaminase; C019969:S-alkylcysteine lyase