title: Purification and amino- and carboxyl-terminal amino acid sequences of alanine-glyoxylate transaminase 1 from human liver.
abstract: In order to confirm the amino acid sequence predicted from the nucleotide sequence of cDNA and also to elucidate the intracellular localization and molecular evolution, human liver alanine-glyoxylate transaminase 1 (AGT1) was purified and subjected to partial amino acid sequence determination, with special attention to posttranslational modification. The enzyme was purified to homogeneity from the 10,000 x g supernatant of human liver homogenate. The purified enzyme showed only a single protein band at about 43 kDa on SDS-PAGE, indicating that it is a homodimer of two identical subunits, because the native enzyme has a molecular mass of about 80 kDa. Both the amino- and carboxyl-terminal peptides of the enzyme were isolated from a cyanogen bromide digest of the S-carboxyl-methylated protein and subjected to amino acid sequence determination. The alpha-amino group of the amino-terminal peptide was shown to be blocked by an acetyl group. The carboxyl-terminal sequence contained a putative N-glycosylation sequence (-Asn-Ala-Thr-), the only one present in the whole molecule, but this sequence was normally determined, indicating that the enzyme is not N-glycosylated. Purdue et al. [J. Cell Biol. 111, 2341-2351 (1990)] have reported that Pro-11, Gly-170, and Ile-340 in normal human AGT1 were replaced by Leu, Arg, and Met, respectively, in a patient with primary hyperoxaluria type 1. We confirmed that residue-11 was Pro. Both the amino- and carboxyl-terminal sequences of the enzyme showed extensive similarity with those of rat liver mitochondrial serine-pyruvate aminotransferase and the small chain of hydrogenase from a thermophilic unicellular cyanobacterium, Synechococcus PCC 6716.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biochemistry, Wakayama Medical College.
authors.forename: I S
authors.initials: IS
authors.lastname: Lee
authors.forename: K
authors.initials: K
authors.lastname: Takio
authors.forename: R
authors.initials: R
authors.lastname: Kido
authors.forename: K
authors.initials: K
authors.lastname: Titani
mesh_terms: D000410:Alanine Transaminase / Q000737:chemistry* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D000818:Animals; D006801:Humans; D008099:Liver / Q000201:enzymology*; D008930:Mitochondria, Liver / Q000201:enzymology; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D051381:Rats; D017386:Sequence Homology, Amino Acid; D000637:Transaminases / Q000737:chemistry
chemical_list: D000596:Amino Acids; D000637:Transaminases; D000410:Alanine Transaminase; C019635:Alanine-glyoxylate transaminase; C022638:serine-pyruvate aminotransferase