The structure of human mitochondrial branched-chain aminotransferase

X-ray crystal structures of three forms of human mitochondrial branched-cha in aminotransferase (BCAT) were solved by molecular-replacement methods, us ing Escherichia coli BCAT as the search model. The enzyme is a homodimer an d the polypeptide chain of each monomer has two domains. The small domain i s composed of residues 1-175 and the large domain is composed of residues 1 76-365. The active site is close to the dimer interface. The 4'-aldehyde of the PLP cofactor is covalently linked to the epsilon -amino group of the a ctive-site lysine, Lys202, via a Schiff-base linkage in two of the structur es. In the third structure, the enzyme is irreversibly inactivated by Tris. The overall fold of the dimer in human mitochondrial BCAT is similar to th e structure of two bacterial enzymes, E. coli BCAT and D-amino acid aminotr ansferase (D-AAT). The residues lining the putative substrate-binding pocke t of human BCAT and D-AAT are completely rearranged to allow catalysis with substrates of opposite stereochemistry. In the case of human mitochondrial branched-chain aminotransferase, a hydrogen-bond interaction between the g uanidinium group of Arg143 in the first monomer with the side-chain hydroxy l of Tyr70 in the second monomer is important in the formation of the subst rate-binding pocket.