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Screening of substrate peptide sequences for tissue-type transglutaminase (TGase 2) using T7 phage cDNA library

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Abstract

Transglutaminase (TGase) is a family of enzymes that catalyzes cross-linking reaction between glutamine- and lysine residue of substrate proteins in several mammalian biological events. Substrate proteins for TGase and their physiological relevance have been still in research, continuously expanding. In this study, we have established a novel screening system that enables identification of cDNA sequence encoding favorable primary structure as a substrate for tissue-type transglutaminase (TGase 2), a multifunctional and ubiquitously expressing isozyme. By the screening, we identified several T7 phage clones that displayed substrate peptides for TGase 2 as a translated product from human brain cDNA library. Among the selected clones, the C-terminal region of IKAP, IkappaB kinase complex associated protein, appeared as a highly reactive substrate sequence for TGase 2. This system will open possibility of rapid identification of substrate sequences for transglutaminases at a genetic level.

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References

  • Anderson SL, Coli R, Daly IW, Kichula EA, Rork MJ, Volpi SA, Ekstein J, Rubin BY (2001) Familial dysautonomia is caused by mutation of the IKAP gene. Am J Hum Genet 68:753–758

    Article  CAS  Google Scholar 

  • Chen Y-T, Hims MM, Shetty RS, Mull J, Liu L, Leyne M, Slaugenhaupt SA (2009) Loss of mouse Ikbkap, a subunit of elongator, leads to transcriptional deficits and embryonic lethality that can be rescued by human IKAKAP. Mol Cell Biol 29:736–744

    Article  CAS  Google Scholar 

  • Cohen L, Henzel WJ, Baeuerie PA (1998) IKAP is a scaffold protein of the IkB kinase complex. Nature 395:292–296

    Article  CAS  Google Scholar 

  • Csosz E, Mesko B, Fésüs L (2009) Transdab wiki: the interactive transglutaminase substrate database on web 2.0 surface. Amino Acids 36:615–617

    Article  CAS  Google Scholar 

  • Eckert RL, Sturniolo MT, Broome AM, Ruse M, Rorke EA (2005) Transglutaminase function in epidermis. J Invest Dermatol 124:481–492

    Article  CAS  Google Scholar 

  • Esposito C, Caputo I (2005) Mammalian transglutaminases: identification of substrates as a key to physiological function and physiological relevance. FEBS J 272:615–631

    Article  CAS  Google Scholar 

  • Facchiano A, Facchiano F (2009) Transglutaminases and their substrates in biology and human diseases: 50 years of growing. Amino Acids 36:599–614

    Article  CAS  Google Scholar 

  • Fésüs L, Piacentini M (2002) Transglutaminase 2: an enigmatic enzyme with diverstic functions. Trends Biochem Sci 27:534–539

    Article  Google Scholar 

  • Griffin M, Casadio R, Bergamini CM (2002) Transglutaminases: nature’s biological glues. Biochem J 368:377–396

    Article  CAS  Google Scholar 

  • Hitomi K (2005) Transglutaminase in skin epidermis. Eur J Dermatol 15:313–319

    CAS  Google Scholar 

  • Hitomi K, Kitamura M, Sugimura Y (2009) Preferred substrate sequences for transglutaminase 2: screening using a phage-displayed peptide library. Amino Acids 36:619–624

    Article  CAS  Google Scholar 

  • Ichikawa A, Ishizaki J, Morita M, Tanaka K, Ikura K (2008) Identification of new amine acceptor protein substrate candidate of transglutaminase in rat liver extract: use of 5-(biotinamido)pentylamine as a probe. Biosci Biotechnol Biochem 72:1056–1062

    Article  CAS  Google Scholar 

  • Ichinose A (2001) Physiopathology and regulation of factor XIII. Thromb Haemost 86:57–65

    CAS  Google Scholar 

  • Ikura K, Kita K, Fujita I, Hashimoto H, Kawabata N (1998) Idetification of amine acceptor protein substrates of transglutaminase in liver extracts: use of 5-(biotinamido)pentylamine as a probe. Arch Biochem Biophys 356:280–286

    Article  CAS  Google Scholar 

  • Jeitner TM, Pinto JT, Krasnikov BF, Horswill M, Cooper AJL (2009) Transglutaminase and neurogeneration. J Neurochem 109(1):160–166

    Article  CAS  Google Scholar 

  • Johansen LD, Naumanen T, Knudsen A, Westerlund N, Gromova I, Junttilla M, Nielsen C, Bottzauw T, Tolkovsky A, Wastermarck J, Coffey ET, Jaattela M, Kallunki T (2008) IKAP localizes to membrane ruffles with filamin A and regulates actin cytoskeleton organization and cell migration. J Cell Sci 121:854–864

    Article  CAS  Google Scholar 

  • Keresztessy Z, Csosz É, Hársfalvi J, Csomós K, Gray J, Lightowlers RN, Lakey JH, Balajthy Z, Fésüs L (2006) Phage display selection of efficient glutamine-donor substrate peptides for transglutaminase 2. Protein Sci 15:2466–2480

    Article  CAS  Google Scholar 

  • Lee KN, Maxwell MD, Patterson MK, Birckbichler PJ, Conway E (1992) Identification of transglutaminase substrates in HT29 colon cancer cells: use of 5-(biotinamido)pentylamine as a transglutaminase-specific probe. Biochim Biophys Acta 1136:12–16

    Article  CAS  Google Scholar 

  • Lorand L, Graham RM (2003) Transglutaminases: crosslinking enzymes with pleiotropic functions. Nat Rev Mol Cell Biol 4:140–156

    Article  CAS  Google Scholar 

  • Morohashi K, Yoshino A, Yoshimori A, Saito S, Tanuma S, Sakaguchi K, Sugawara F (2005) Identification of a drug target motif: an anti-tumor drug NK109 interacts with a PNxxxxP. Biochem Pharmacol 70:37–46

    Article  CAS  Google Scholar 

  • Orru S, Caputo I, D’Amato A, Ruoppolo M, Esposito C (2003) Proteomics identification of acyl-acceptor and acyl-donor substrates for transglutaminase in a human intestinal epithelial cell line. J Biol Chem 278:31766–31773

    Article  CAS  Google Scholar 

  • Slaugenhaupt SA, Blumenfeld A, Gill SP, Leyne M, Mull J, Cuajungco MP, Liebert CB, Chadwick B, Idelson M, Reznik L, Robbins CM, Makalowska I, Brownstein M, Krappman D, Scheidereit C, Maayan C, Axelrod FB, Gusella JF (2001) Tissue-specific expression of a splicing mutation in the IKABKAP gene causes familial dysautonomia. Am J Hum Genet 68:598–605

    Article  CAS  Google Scholar 

  • Sollid LM (2002) Coeliac disease: dissecting a complex inflammatory disorder. Nat Rev Immunol 2:647–655

    Article  CAS  Google Scholar 

  • Sugimura Y, Hosono M, Wada F, Yoshimura T, Maki M, Hitomi K (2006) Screening for the preferred substrate sequence of transglutaminase using a phage-displayed peptide library: identification of peptide substrates for TGase 2 and factor XIIIa. J Biol Chem 281:17699–17706

    Article  CAS  Google Scholar 

  • Sugimura Y, Hosono M, Kitamura M, Tsuda T, Yamanishi K, Maki M, Hitomi K (2008a) Identification of preferred substrate sequences for transglutaminase 1-development of a novel peptide that can efficiently detect cross-linking enzyme activity in the skin. FEBS J 275:5667–5677

    CAS  Google Scholar 

  • Sugimura Y, Yokoyma K, Nio N, Maki M, Hitomi K (2008b) Identification of preferred substrate sequences of microbial transglutaminase from Streptomyces mobaraensis using a phage-displayed peptide library. Arch Biochem Biophys 477:379–383

    Article  CAS  Google Scholar 

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Acknowledgments

We greatly appreciate Dr. Masatoshi Maki and Dr. Hideki Shibata in our laboratory for providing valuable suggestions. This work was supported by a Grant-in-Aid for Scientific Research on Innovative Areas (No. 20200072) from the Ministry of Education, Sports, Science and Technology (MEXT, Japan) (to K. H.) and also Grant-in-Aid for Young Scientists Research (No. 186701) from the Japan Society for the Promotion of Science (JSPS) (to Y. S.).

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  1. Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, 464-8601, Japan

    Yoshiaki Sugimura, Hiroyuki Yamashita & Kiyotaka Hitomi

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  1. Yoshiaki Sugimura
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  2. Hiroyuki Yamashita
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  3. Kiyotaka Hitomi
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Correspondence to Kiyotaka Hitomi.

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Sugimura, Y., Yamashita, H. & Hitomi, K. Screening of substrate peptide sequences for tissue-type transglutaminase (TGase 2) using T7 phage cDNA library. Cytotechnology 63, 111–118 (2011). https://doi.org/10.1007/s10616-010-9308-7

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  • DOI: https://doi.org/10.1007/s10616-010-9308-7

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