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Crystal structure of an oxidized mutant of human mitochondrial branched‐chain aminotransferase
Author(s) -
Herbert Darius,
Gibbs Synphane,
Riddick Alexys,
Conway Myra,
Dong Ming
Publication year - 2020
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x19016480
Subject(s) - transamination , active site , stereochemistry , cysteine , chemistry , valine , mutagenesis , leucine , crystal structure , active center , amino acid , isoleucine , mutant , side chain , site directed mutagenesis , protein structure , biochemistry , crystallography , enzyme , organic chemistry , gene , polymer
This study presents the crystal structure of a thiol variant of the human mitochondrial branched‐chain aminotransferase protein. Human branched‐chain aminotransferase (hBCAT) catalyzes the transamination of the branched‐chain amino acids leucine, valine and isoleucine and α‐ketoglutarate to their respective α‐keto acids and glutamate. hBCAT activity is regulated by a C XX C center located approximately 10 Å 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. Site‐directed mutagenesis 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. The crystal structure of the oxidized form of the C318A variant was used to better understand the contributions of the individual cysteines and their oxidation states. The structure reveals the modified C XX C 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. Moreover, the structure reveals conformational differences in the N‐terminal and inter‐domain region that may correlate with the inactivated state of the C XX C center.