Abstract
γ-Glutamylamine cyclotransferase (gGACT) catalyzes the intramolecular cyclization of a variety of l-γ-glutamylamines producing 5-oxo-l-proline and free amines. Its substrate specificity implicates it in the downstream metabolism of transglutaminase products, and is distinct from that of γ-glutamyl cyclotransferase which acts on l-γ-glutamyl amino acids. To elucidate the mechanism by which gGACT distinguishes between l-γ-glutamylamine and amino acid substrates, the specificity of the rabbit kidney enzyme for the amide region of substrates was probed through the kinetic analysis of a series of l-γ-glutamylamines. The isodipeptide N ɛ-(l-γ-glutamyl)-l-lysine 1 was used as a reference. The kinetic constants of the l-γ-glutamyl derivative of n-butylamine 7, were nearly identical to those of 1. Introduction of a methyl or carboxylate group on the carbon adjacent to the side-chain amide nitrogen in l-γ-glutamylamine substrates resulted in a dramatic decrease in substrate properties for gGACT thus providing an explanation of why gGACT does not act on l-γ-glutamyl amino acids except for l-γ-glutamylglycine. Placement of substituents on carbons further removed from the side-chain amide nitrogen in l-γ-glutamylamines restored activity for gGACT, and l-γ-glutamylneohexylamine 19 had a higher specificity constant (k cat /K m) than 1. gGACT did not exhibit any stereospecificity in the amide region of l-γ-glutamylamine substrates. In addition, analogues (26–30) with heteroatom substitutions for the γ methylene position of the l-γ-glutamyl moiety were examined. Several thiocarbamoyl derivatives of l-cysteine (28–30) were excellent substrates for gGACT.
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References
Beninati S, Bergamini CM, Piacentini M (2009) An overview of the first 50 years of transglutaminase research. Amino Acids 36:591–598
Beninati S, Piacentini M, Argento-Cerù MP, Russo-Caia S, Autuori F (1985) Presence of di- and polyamines covalently bound to protein in rat liver. Biochim Biophys Acta 841(1):120–126
Danson JW, Trawick ML, Cooper AJL (2002) Spectrophotometric assays for l-lysine alpha-oxidase and gamma-glutamylamine cyclotransferase. Anal Biochem 303:120–130
Davies PJ, Murtaugh MP, Moore WT Jr, Johnson GS, Lucas D (1985) Retinoic acid-induced expression of tissue transglutaminase in human promyelocytic leukemia (HL-60) cells. J Biol Chem 260:5166–5174
Ferraz de Souza W, Kambe N, Sonoda N (1996) Theoretical study of structures and internal rotations of methyl N, N-dimethylcarbamate and its sulfur, selenium, and tellurium homologs (Me2NC(O)YMe, Y=O, S, Se, Te). J Phys Org Chem 9(3):179–186
Facchiano A, Facchiano F (2009) Transglutaminases and their substrates in biology and human diseases: 50 years of growing. Amino Acids 36:599–614
Fesus L, Davies PJ, Piacentini M (1991) Apoptosis: molecular mechanisms in programmed cell death. Eur J Cell Biol 56(2):170–177
Fink ML, Chung SI, Folk JE (1980) gamma-Glutamylamine cyclotransferase: specificity toward epsilon-(l-gamma-glutamyl)-l-lysine and related compounds. Proc Natl Acad Sci USA 77:4564–4568
Fink ML, Folk JE (1981a) γ-Glutamylamine cyclotransferase: an enzyme involved in the catabolism of ε-(γ-glutamyl)lysine and other γ-glutamylamines. Mol Cell Biochem 38:59–67
Fink ML, Folk JE (1981b) γ-Glutamylpolyamines: implications in the degradation of polyamine-protein conjugates. In Caldarera CM et al (eds) Advances in polyamine research, vol 3. Raven Press, New York, pp 187–196
Fink ML, Folk JE (1983) gamma-Glutamylamine cyclotransferase (rabbit kidney). In: Tabor H, Tabor CW (eds) Polyamines. Methods enzymology, vol 94. Academic Press, New York, pp 347–351
Folk JE, Finlayson JS (1977) The epsilon-(gamma-glutamyl)lysine crosslink and the catalytic role of transglutaminases. Adv Protein Chem 31:1–133
Folk JE (1980) Transglutaminases. Ann Rev Biochem 49:517–531
Folk JE, Park MH, Chung SI, Schrode J, Lester EP, Cooper HL (1980) Polyamines as physiological substrates for transglutaminases. J Biol Chem 255:3695–3700
Oakley AJ, Yamada T, Liu D, Coggan M, Clark AG, Board PG (2008) The identification and structural characterization of C7orf24 as γ-glutamyl cyclotransferase: an essential enzyme in the γ-glutamyl cycle. J Biol Chem 283:22031–22042
Oakley AJ, Coggan M, Clark AG, Board PG (2010) Identification and characterization of γ-glutamylamine cyclotransferase, an enzyme responsible for γ-glutamyl-ε-lysine catabolism. J Biol Chem 285:9642–9648
Orlowski M, Meister A (1973) The γ-glutamyl cyclotransferase. Distribution, isoenzymic forms, and specificity. J Biol Chem 248(8):2836–2844
Orlowski M, Richman PG, Meister A (1969) Isolation and properties of γ-l-glutamylcyclotransferase from human brain. Biochemistry 8(3):1048–1055
Waszkowycz B, Hillier IH, Gensmantel N, Payling DW (1991) Combined quantum mechanical-molecular mechanical study of catalysis by the enzyme phospholipase A2: an investigation of the potential energy surface for amide hydrolysis. J Chem Soc Perkin Trans 2; Phys Org Chem (1972–1999) (12):2025–2032
Acknowledgments
The authors are grateful for the contribution of the late Dr. John E. Folk who synthesized many of the compounds used in this study. We express our appreciation to Mr. Gustavo E. Chavarria for expert assistance in the preparation of this manuscript. This work was supported by the University Research Committee, Baylor University.
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Dedicated to the late Dr. John E. Folk, who synthesized many of the compounds used in this study, for his long-standing collaboration, and his unfailing encouragement.
Mary Lynn Trawick was formerly Mary Lynn Fink.
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Bowser, T.E., Trawick, M.L. Probing the specificity of gamma-glutamylamine cyclotransferase: an enzyme involved in the metabolism of transglutaminase-catalyzed protein crosslinks. Amino Acids 44, 143–150 (2013). https://doi.org/10.1007/s00726-011-1153-2
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DOI: https://doi.org/10.1007/s00726-011-1153-2