Determination of amino- and carboxyl-terminal sequences of guinea pig liver transglutaminase: evidence for amino-terminal processing.
Transglutaminases (EC 2.3.2.13) catalyze the formation of epsilon-(gamma-glutamyl)lysine cross-links and the substitution of a variety of primary amines for the gamma-carboxamide groups of protein-bound glutaminyl residues. These enzymes are involved in many biological phenomena. In this study, the amino- and carboxyl-terminal sequences of guinea pig liver transglutaminase were identified by sequence analysis to determine whether this enzyme is processed posttranslationally at its terminal regions. Two peptides, believed to contain the amino-terminal sequences of transglutaminase, were isolated from the Pronase digest of the enzyme protein with SP-Sephadex C-25 column chromatography and reverse-phase HPLC. Analyses (amino acid analysis, sequencing after the treatment with an acylamino-acid-releasing enzyme, and fast atom bombardment mass spectrometry) of these peptides indicated that the amino-terminal structure of this enzyme is acetylAla-Glu-Asp-Leu-Ile-Leu-Glu. The candidate for the carboxyl-terminal peptide in the trypsin digest of enzyme was isolated from the unadsorbed fraction of affinity chromatography with anhydrotrypsin agarose gel. The peptide was found to be Asn-Val-Ile-Ile-Gly-Pro-Ala. Both the terminal sequences were completely consistent with those predicted from the cDNA sequence [Ikura, K., Nasu, T., Yokota, H., Tsuchiya, Y., Sasaki, R., & Chiba, H. (1988) Biochemistry 27, 2898-2905]. These results indicated that the amino-terminal processing occurred after or in the course of translation of this enzyme, namely, removal of the initiator methionine and a subsequent acetylation of the alanine residue adjacent to the methionine. Our results did not indicate carboxyl-terminal processing of guinea pig liver transglutaminase.
Ikura
K
K
Department of Food Science and Technology, Faculty of Agriculture, Kyoto University, Japan.
Yokota
H
H
Sasaki
R
R
Chiba
H
H
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid; D006168:Guinea Pigs; D008099:Liver / Q000201:enzymology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D011499:Protein Processing, Post-Translational; D011503:Transglutaminases / Q000235:genetics* / Q000378:metabolism*
D000596:Amino Acids; D011503:Transglutaminases