SEQUENCE ANALYSIS AND STRUCTURAL CHARACTERIZATION OF TISSUE TRANSGLUTAMINASE 2(TG2) BY IN SILICO APPROACH
Objective: TG2 is multifunctional protein. The up regulation leads into different pathological disorders. The objective of the present study was the prediction of a structural feature of TG2 (Tissue transglutaminase) protein with in silico approach.
Methods: In this study, we have investigated the structural feature of TG2 by using various biological databases (uniprot, NCBI, Pfam) and online tools such as BLAST, PDBsum, protoparam tools.
Results: The predicted structure of TG2 protein has shown that amino acid residues conserved throughout the sequence in selected mammals. During the course of evolution, mammalian TG2 protein is orthologus; human TG2 shares its characters with chimpanzee while mice and rat were closely related to each other. This protein was mainly cytosolic but also present in other cell organalles. It has four catalytic domains and three active sites with multiple metal binding domain specifically for calcium. The pI value was 5.11, GRAVY-0.283. The phosphorylation sites were present at serine and threonine. The structure was a monomer with 14 alpha helices and 9 sheets. Ramachandran plot showed about 98% residues in the favoured region.
Conclusion: Collectively, these data suggest that the predicted TG2 protein may act as a good therapeutic target. Targeting phosphorylation sites may help in down regulation of TG2. The modelled protein can be used for further work.
2. Achyuthan KE, Greenberg CS. Identification of a guanosine triphosphate-binding site on guinea pig liver transglutaminase. The role of GTP and calcium ions in modulating activity. J Biol Chem 1987 5;262:1901-6.
3. Mangala LS, Mehta K. Tissue transglutaminase (TG2) in cancer biology. Prog Exp Tumor Res 2005;38:125â€“38.
4. Csosz E, Bagossi P, Nagy Z, Dosztanyi Z, Simon I, Fesus L. Substrate preference of transglutaminase 2 revealed by logistic regression analysis and intrinsic disorder examination. J Med Bacteriol 2008;383:390-2.
5. Chhabra A, Verma A, Mehta K. Tissue transglutaminase promotes or suppresses tumors depending on cell context. Anticancer Res 2009;29:1909-19.
6. Iismaa SE, Mearns BM, Lorand L, Graham RM. Transglutaminases and disease: lessons from genetically engineered mouse models and inherited disorders. Phys Res 2009;89:991-1023.
7. Antonyak MA, Li B, Regan AD, Feng Q, Dusaban SS, Cerione RA. Tissue transglutaminase is an essential participant in the epidermal growth factor-stimulated signalling pathway leading to cancer cell migration and invasion. J Biol Chem 2009;284:17914-25.
8. Nanda N, Iismaa SE, Owens WA, Husain A, Mackay F, Graham RM. Targeted inactivation of Gh/tissue transglutaminase II. J Biol Chem 2001;276:20673-8.
9. Hasegawa G, Motoi SU, Ichikawa Y, Ohtsuka T, Kumagai S, Kikuchi M, et al. A novel function of tissue-type transglutaminase: protein disulphide isomerase. Biochemical J 2003;373:793-3.
10. Mishra S, Murphy LJ. The p53 oncoprotein is a substrate for tissue transglutaminase kinase activity. Biochem Biophys Res Commu 2006;339:726-30.
11. Wang Y, Ande SR, Mishra S. Phosphorylation of transglutaminase 2 (TG2) at serine-216 has a role in TG2 mediated activation of nuclear factor-kappa B and in the downregulation of PTEN. BMC Cancer 2012;12:277.
12. Mishra S, Saleh A, Espino PS, Davie JR, Murphy LJ. Phosphorylation of histones by tissue transglutaminase. J Biol Chem 2006;281:5532-8.
13. Mishra S, Murphy LJ. Phosphorylation of transglutaminase 2 by PKA at Ser216 creates 14-3-3 binding sites. Biochem Biophys Res Commun 2006;347:1166-70.
14. Park H, Park ES, Lee HS, Yun HY, Kwon NS, Baek KJ. A distinct characteristics of GÎ±h (transglutaminase II) by compartment: GTPase and transglutaminase activities. Biochem Biophys Res Commun 2001;284:496-500.
15. Park D, Choi SS, Ha KS. Transglutaminase 2: a multi-functional protein in multiple subcellular compartments. Amino Acids 2010;39:619-31.
16. Pinkas DM, Strop P, Brunger AT, Khosla C. Transglutaminase 2 undergoes a large conformational change upon activation. PLoS Biol 2007;5:327.
17. Kanchan K, Fuxreiter M, FÃ©sÃ¼s L. Physiological, pathological, and structural implications of non-enzymatic protein-protein interactions of the multifunctional human transglutaminase 2. Cell Mol Life Sci 2015;72:3009-35.
18. Liu S, Cerione RA, Clardy J. Structural basis for the guanine nucleotide-binding activity of tissue transglutaminase and its regulation of transamidation activity. Proceedings Nat Acad Sci 2002;99:2743-7.
19. Thiery JP, Acloque H, Huang RY, Nieto MA. Epithelial-mesenchymal transitions in development and disease. Cell 2009;139:871-90.
20. Hoffner G, Djian P. Transglutaminase and diseases of the central nervous system. Front Biosci 2005;10:3078-92.
21. Lai TS, Liu Y, Tucker T, Daniel KR, Sane DC, Toone E, et al. Identification of chemical inhibitors to human tissue transglutaminase by screening existing drug libraries. Chem Biol 2008;15:969-78.
22. Jang TH, Lee DS, Choi K, Jeong EM, Kim IG, Kim YW, et al. Crystal structure of transglutaminase 2 with GTP complex and amino acid sequence evidence of the evolution of GTP binding site. PLoS One 2014;9:7005.
23. Han BG, Cho JW, Cho YD, Jeong KC, Kim SY, Lee BI. Crystal structure of human transglutaminase 2 in complex with adenosine triphosphate. Int J Biomater 2010;47:190-5.
24. Bergamini CM, Dondi A, Lanzara V, Squerzanti M, Cervellati C, Montin K, et al. Thermodynamics of binding of regulatory ligands to tissue transglutaminase. Amino Acids 2010;39:297-304.
25. McWilliam H, Li W, Uludag M, Squizzato S, Park YM, Buso N, et al. Analysis tool web services from the EMBL-EBI. Nucleic Acids Res 2013;41:597-600.
26. Finn RD, Coggill P, Eberhardt RY, Eddy SR, Mistry J, Mitchell AL, et al. The pfam protein familiesâ€™ database: towards a more sustainable future. Nucleic Acids Res 2016;44:279-85.
27. Boratyn GM, Camacho C, Cooper PS, Coulouris G, Fong A, Ma N, et al. BLAST: a more efficient report with usability improvements. Nucleic Acids Res 2013;41:29-33.
28. Yu CS, Lin CJ, Hwang JK. Predicting subcellular localization of proteins for Gram-negative bacteria by support vector machines based on nâ€peptide compositions. Pro Sci 2004;13:1402-6.
29. Petersen TN, Brunak S, von Heijne G, Nielsen H. Signal P 4.0: discriminating signal peptides from transmembrane regions. Nat Methods 2011;8:785-6.
30. Gasteiger E, Hoogland C, Gattiker A, Duvaud SE, Wilkins MR, Appel RD, et al. Protein identification and analysis tools on the ExPASy server. Humana Press; 2005.
31. Blom N, Gammeltoft S, Brunak S. Sequence and structure-based prediction of eukaryotic protein phosphorylation sites. J Mol Biol 1999;294:1351-62.
32. Petersen B, Petersen TN, Andersen P, Nielsen M, Lundegaard C. A generic method for assignment of reliability scores applied to solvent accessibility predictions. BMC Struct Biol 2009;9:51.
33. Possner DD, Claesson M, Guy JE. Structure of the glycosyltransferase Ktr4p from saccharomyces cerevisiae. PloS One 2015;10:0136239. https://doi.org/10.1371/journal.pone. 0136239
34. Hintze BJ, Lewis SM, Richardson JS, Richardson DC. Molprobity's ultimate rotamerâ€library distributions for model validation. Proteins: Struct Funct Bioinf 2016;84:1177-89.
35. Biasini M, Bienert S, Waterhouse A, Arnold K, Studer G, Schmidt T, et al. SWISS-MODEL: modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Res 2014;42:W252-8.
36. Colovos C, Yeates TO. Verification of protein structures: patterns of nonbonded atomic interactions. Proteins Sci 1993;2:1511-9.
37. Henderson KN, Tye-Din JA, Reid HH, Chen Z, Borg NA, Beissbarth T, et al. A structural and immunological basis for the role of human leukocyte antigen DQ8 in celiac disease. Immunity 2007;27:23-34.
38. Mangala LS, Mehta K. Tissue transglutaminase (TG2) in cancer biology. In: Transglutaminases. Karger Pub; 2005. p. 125-38.
39. Sohn J, Kim TI, Yoon YH, Kim JY, Kim SY. Novel transglutaminase inhibitors reverse the inflammation of allergic conjunctivitis. J Clin Inves 2003;111:121-8.
40. Siegel M, Khosla C. Transglutaminase 2 inhibitors and their therapeutic role in disease states. Pharmacol Ther 2007;115:232-45.
41. Mariani P, Carsughi F, Spinozzi F, Romanzetti S, Meier G, Casadio R, et al. Ligand-induced conformational changes in tissue transglutaminase: Monte Carlo analysis of small-angle scattering data. Biophys J 2000;78:3240â€“51.
42. Milakovic T, Tucholski J, McCoy E, Johnson GV. Intracellular localization and activity state of tissue transglutaminase differentially impacts cell death. J Biol Chem 2004;279: 8715-22.
43. Bergamini CM, Signorini M. Purification of testicular transglutaminase by hydrophobic chromatography on phenyl-sepharose. Biochem Int 1992;27:557-65.
44. Huang L, Haylor JL, Hau Z, Jones RA, Vickers ME, Wagner B, et al. Transglutaminase inhibition ameliorates experimental diabetic nephropathy. Kidney Int 2009;76:383-94.
45. Johnson TS, El-Koraie AF, Skill NJ, Baddour NM, El Nahas AM, Njloma M, et al. Tissue transglutaminase and the progression of human renal scarring. J Am Soc Nephrol 2003;14:2052-62.
46. Chica RA, Gagnon P, Keillor JW, Pelletier JN. Tissue transglutaminase acylation: the proposed role of conserved active site Tyr and Trp residues revealed by molecular modeling of peptide substrate binding. Proteins Sci 2004;13:979-91.
47. Wang Y, Ande SR, Mishra S. Overexpression of phospho mutant forms of transglutaminase 2 downregulates epidermal growth factor receptor. Biochem Biophys Res Commun 2012;417:251â€“5.
48. Begg GE, Holman SR, Stokes PH, Matthews JM, Graham RM, et al. Mutation of a critical arginine in the GTP-binding site of transglutaminase 2 disinhibits intracellular cross-linking activity. J Biol Chem 2006;281:12603â€“9.
49. Stamnaes J, Pinkas DM, Fleckenstein B, Khosla C, Sollid LM. Redox regulation of transglutaminase 2 activity. J Biol Chem 2010;285:25402â€“9.
50. Umashankar V, Gurunathan S. Drug discovery: an appraisal. Int J Pharm Pharm Sci 2015;7:59-66.