Protein structure of pig liver 4‐aminobutyrate aminotransferase and comparison with a cDNA‐deduced sequence

Abstract

The amino acid sequence of pig liver 4‐aminobutyrate aminotransferase has been determined by gas‐phase sequencing of proteolytically derived peptide fragments. The sequence differs substantially from that predicted for the same enzyme on the basis of the sequence of cDNA derived from pig brain in recently published work [Kwon, O., Park, J. & Churchich, J. E. (1992) J. Biol. Chem. 267, 7215–7216]. Apart from a few minor differences, the two sequences are completely different in the segment of protein comprising the 36 residues at positions 107–142. Insertion of a cytosine between bases 402 and 403 in the cDNA sequence, together with deletion of the guanine at position 510, results in a DNA sequence which predicts exactly the amino acid sequence determined by peptide analysis in the present work. The mammalian enzyme has approximately 44% sequence identity with the same enzyme from two unicellular eukaryotes (Saccharomyces cerevisiae, Aspergillus nidulans) and 22% identity with that from Escherichia coli.

Primary Structure and Tissue Distribution of Human 4‐Aminobutyrate Aminotransferase

“…More than 90% of the sequence information vvas obtained by purification and analysis of an almost complete set of tryptic peptides that were overlapped in part by peptides obtained from CNBr and Asp-Pro bond cleavages. Due to the very high extent of similarity between human and pig 4-Abu aminotransferase (vide infru), the available primary structure of the porcine enzyme [13] was useful for easy ordering of the fragments of human 4-Abu aminotransferase reconstructed from the above-mentioned sets of peptides.…”

“…One of these drugs is vigabatrin, a vinylic substrate analog [9, 101. We have studied in detail the mechanism of inactivation of 4-Abu aminotransferase by vigabatrin [11], now successfully used in the treatment of some forms of epilepsy [ 121, using the enzyme prepared from pig liver as a model ; the primary structure of this enzyme was determined in our laboratory [13]. In an earlier study, we also investigated the stoichiometry and stability of the adduct formed between vigabatrin and human liver 4-Abu aminotransferase [14].…”

The 2011 E. B. Hershberg Award for Important Discoveries in Medicinally Active Substances: (1S,3S)-3-Amino-4-difluoromethylenyl-1-cyclopentanoic Acid (CPP-115), a GABA Aminotransferase Inactivator and New Treatment for Drug Addiction and Infantile Spasms

“…The primary sequence of GABA-AT has been deduced from the cDNA of pig brain 42 and from peptide fragments of the pig liver enzyme. 43 In 1999 the X-ray crystal structure of pig liver GABA-AT was reported to 3.0 Å resolution by the Schirmer group in Basel. 44 A 1.9 Å resolution crystal structure with one of our inactivators bound also has been reported.…”

Structures of γ-Aminobutyric Acid (GABA) Aminotransferase, a Pyridoxal 5′-Phosphate, and [2Fe-2S] Cluster-containing Enzyme, Complexed with γ-Ethynyl-GABA and with the Antiepilepsy Drug Vigabatrin

“…GABA-AT is a homodimer with each subunit containing an active-site PLP covalently bound to Lys-329 via a Schiff base. The primary sequence of GABA-AT has been deduced from the cDNA of pig brain (3) and from peptide fragments of the pig liver enzyme (4). Together with the structurally well characterized enzymes ornithine aminotransferase (OAT) and dialkylglycine decarboxylase (DGD), GABA-AT belongs to the subfamily II of the ␣-family of PLP-dependent enzymes (5).…”