7LON

Ornithine Aminotransferase (OAT) cocrystallized with its inactivator - (1S,3S)-3-amino-4-(difluoromethylene)cyclohexene-1-carboxylic acid

Summary for 7LON

• Entry DOI: 10.2210/pdb7lon/pdb
• Descriptor: Ornithine aminotransferase, mitochondrial, THREONINE, (1R,3S,4R)-3-[({3-hydroxy-2-methyl-5-[(phosphonooxy)methyl]pyridin-4-yl}methyl)amino]-4-methylcyclohexane-1-carboxylic acid, ... (4 entities in total)
• Functional Keywords: ornithine aminotransferase, oat, aminotransferase, inactivator, inhibitor, transferase
• Biological source: Homo sapiens (Human)
• Total number of polymer chains: 3
• Total formula weight: 136103.38

Authors

Butrin, A.,Zhu, W.,Liu, D.,Silverman, R. (deposition date: 2021-02-10, release date: 2021-08-25, Last modification date: 2024-11-06)

Primary citation

Zhu, W.,Doubleday, P.F.,Butrin, A.,Weerawarna, P.M.,Melani, R.D.,Catlin, D.S.,Dwight, T.A.,Liu, D.,Kelleher, N.L.,Silverman, R.B.
Remarkable and Unexpected Mechanism for ( S )-3-Amino-4-(difluoromethylenyl)cyclohex-1-ene-1-carboxylic Acid as a Selective Inactivator of Human Ornithine Aminotransferase.
J.Am.Chem.Soc., 143:8193-8207, 2021

PubMed Abstract: Human ornithine aminotransferase (OAT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that was recently found to play an important role in the metabolic reprogramming of hepatocellular carcinoma (HCC) via the proline and glutamine metabolic pathways. The selective inhibition of OAT by compound exhibited potent antitumor activity. Inspired by the discovery of the aminotransferase inactivator (1,3)-3-amino-4-(difluoromethylene)cyclopentane-1-carboxylic acid (), we rationally designed, synthesized, and evaluated a series of six-membered-ring analogs. Among them, was identified as a new selective OAT inactivator, which demonstrated a potency 22× greater than that of . Three different types of protein mass spectrometry approaches and two crystallographic approaches were employed to identify the structure of OAT- and the formation of a remarkable final adduct () in the active site. These spectral studies reveal an enzyme complex heretofore not observed in a PLP-dependent enzyme, which has covalent bonds to two nearby residues. Crystal soaking experiments and molecular dynamics simulations were carried out to identify the structure of the active-site intermediate and elucidate the order of the two covalent bonds that formed, leading to . The initial covalent reaction of the activated warhead occurs with *Thr322 from the second subunit, followed by a subsequent nucleophilic attack by the catalytic residue Lys292. The turnover mechanism of by OAT was supported by a mass spectrometric analysis of metabolites and fluoride ion release experiments. This novel mechanism for OAT with will contribute to the further rational design of selective inactivators and an understanding of potential inactivation mechanisms by aminotransferases.

PubMed: 34014654
DOI: 10.1021/jacs.1c03572

Experimental method

X-RAY DIFFRACTION (1.95 Å)