Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Twin-arginine signal peptide attributes effective display of CD147 to filamentous phage

  • 175 Accesses

  • 11 Citations

Abstract

A novel phagemid (pTat8) was constructed in this study to improve the quality of a molecule displayed on filamentous phage. The twin-arginine translocation (Tat) pathway was chosen for transporting and integrating a CD147 molecule into a phage particle via gpVIII. The parent vector pComb8-CD147Ex was modified by substituting a Sec signal sequence (PelB) with a twin-arginine signal sequence from trimethylamine N-oxide reductase (TorA). The characteristics of the CD147 displayed on the phage particle were evaluated by Sandwich ELISA and Western immunoblotting. A Tat-dependent leader was found to be superior to the Sec leader for the phage display of CD147. Our findings further support the involvement of an Escherichia coli Tat translocase in mediating the integration of a hydrophobic transmembrane protein into the inner membrane. This modified phagemid will be useful in phage display technique when the correctly folded structure is required (i.e., antibody libraries and ligand–receptor tracing).

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  1. Barbas CF, Burton DR, Scott JK, Silverman GJ (2001) Phage display: a laboratory manual. Cold Spring Harbor Laboratory Press, New York

  2. Bass S, Greene R, Wells JA (1990) Hormone phage: an enrichment method for variant proteins with altered binding properties. Proteins 8:309–314

  3. Berks BC (1996) A common export pathway for proteins binding complex redox cofactors? Mol Microbiol 22:393–404

  4. Berks BC, Sargent F, Palmer T (2000) The Tat protein export pathway. Mol Microbiol 35:260–274

  5. Biswas C, Zhang Y, DeCastro R, Guo H, Nakamura T, Kataoka H, Nabeshima K (1995) The human tumor cell-derived collagenase stimulatory factor (renamed EMMPRIN) is a member of the immunoglobulin superfamily. Cancer Res 55:434–439

  6. Bothmann H, Pluckthun A (1998) Selection for a periplasmic factor improving phage display and functional periplasmic expression. Nat Biotechnol 16:376–380

  7. Bruser T, Sanders C (2003) An alternative model of the twin arginine translocation system. Microbiol Res 158:7–17

  8. Chappel JA, He M, Kang AS (1998) Modulation of antibody display on M13 filamentous phage. J Immunol Methods 221:25–34

  9. Danese PN, Silhavy TJ (1998) Targeting and assembly of periplasmic and outer-membrane proteins in Escherichia coli. Annu Rev Genet 32:59–94

  10. DeLisa MP, Lee P, Palmer T, Georgiou G (2004) Phage shock protein PspA of Escherichia coli relieves saturation of protein export via the Tat pathway. J Bacteriol 186:366–373

  11. Endemann H, Model P (1995) Location of filamentous phage minor coat proteins in phage and in infected cells. J Mol Biol 250:496–506

  12. Hatzixanthis K, Palmer T, Sargent F (2003) A subset of bacterial inner membrane proteins integrated by the twin-arginine translocase. Mol Microbiol 49:1377–1390

  13. Heyd B, Pecorari F, Collinet B, Adjadj E, Desmadril M, Minard P (2003) In vitro evolution of the binding specificity of neocarzinostatin, an enediyne-binding chromoprotein. Biochemistry 42:5674–5683

  14. Hoogenboom HR, Griffiths AD, Johnson KS, Chiswell DJ, Hudson P, Winter G (1991) Multi-subunit proteins on the surface of filamentous phage: methodologies for displaying antibody (Fab) heavy and light chains. Nucleic Acids Res 19:4133–4137

  15. Intasai N, Arooncharus P, Kasinrerk W, Tayapiwatana C (2003) Construction of high-density display of CD147 ectodomain on VCSM13 phage via gpVIII: effects of temperature, IPTG, and helper phage infection-period. Protein Expr Purif 32:323–331

  16. Ize B, Stanley NR, Buchanan G, Palmer T (2003) Role of the Escherichia coli Tat pathway in outer membrane integrity. Mol Microbiol 48:1183–1193

  17. Kasinrerk W, Fiebiger E, Stefanova I, Baumruker T, Knapp W, Stockinger H (1992) Human leukocyte activation antigen M6, a member of the Ig superfamily, is the species homologue of rat OX-47, mouse basigin, and chicken HT7 molecule. J Immunol 149:847–854

  18. Kasinrerk W, Tokrasinwit N, Phunpae P (1999) CD147 monoclonal antibodies induce homotypic cell aggregation of monocytic cell line U937 via LFA-1/ICAM-1 pathway. Immunology 96:184–192

  19. Manosroi J, Tayapiwatana C, Gotz F, Werner RG, Manosroi A (2001) Secretion of active recombinant human tissue plasminogen activator derivatives in Escherichia coli. Appl Environ Microbiol 67:2657–2664

  20. Marciano DK, Russel M, Simon SM (1999) An aqueous channel for filamentous phage export. Science 284:1516–1519

  21. Mitraki A, Fane B, Haase-Pettingell C, Sturtevant J, King J (1991) Global suppression of protein folding defects and inclusion body formation. Science 253:54–58

  22. Paschke M, Hohne W (2005) A twin-arginine translocation (Tat)-mediated phage display system. Gene 350:79–88

  23. Pugsley AP (1993) The complete general secretory pathway in gram-negative bacteria. Microbiol Rev 57:50–108

  24. Sambasivarao D, Dawson HA, Zhang G, Shaw G, Hu J, Weiner JH (2001) Investigation of Escherichia coli dimethyl sulfoxide reductase assembly and processing in strains defective for the sec-independent protein translocation system membrane targeting and translocation. J Biol Chem 276:20167–20174

  25. Santini CL, Ize B, Chanal A, Muller M, Giordano G, Wu LF (1998) A novel sec-independent periplasmic protein translocation pathway in Escherichia coli. EMBO J 17:101–112

  26. Sargent F, Bogsch EG, Stanley NR, Wexler M, Robinson C, Berks BC, Palmer T (1998) Overlapping functions of components of a bacterial Sec-independent protein export pathway. EMBO J 17:3640–3650

  27. Sidhu SS, Weiss GA, Wells JA (2000) High copy display of large proteins on phage for functional selections. J Mol Biol 296:487–495

  28. Soltes G, Barker H, Marmai K, Pun E, Yuen A, Wiersma EJ (2003) A new helper phage and phagemid vector system improves viral display of antibody Fab fragments and avoids propagation of insert-less virions. J Immunol Methods 274:233–244

  29. Stanley NR, Findlay K, Berks BC, Palmer T (2001) Escherichia coli strains blocked in Tat-dependent protein export exhibit pleiotropic defects in the cell envelope. J Bacteriol 183:139–144

  30. Tayapiwatana C, Kasinrerk W (2002) Construction and characterization of phage-displayed leukocyte surface molecule CD99. Appl Microbiol Biotechnol 60:336–341

  31. Thomas JD, Daniel RA, Errington J, Robinson C (2001) Export of active green fluorescent protein to the periplasm by the twin-arginine translocase (Tat) pathway in Escherichia coli. Mol Microbiol 39:47–53

  32. Vallejo LF, Rinas U (2004) Strategies for the recovery of active proteins through refolding of bacterial inclusion body proteins. Microb Cell Fact 3:11

  33. Weiner JH, Bilous PT, Shaw GM, Lubitz SP, Frost L, Thomas GH, Cole JA, Turner RJ (1998). A novel and ubiquitous system for membrane targeting and secretion of cofactor-containing proteins. Cell 93:93–101

Download references

Acknowledgement

We are grateful to Dr. Nicole Ngo (The Perinatal HIV Prevention Trial Group, Chiang Mai, Thailand) for cooperation in determining the inserted fragment in pTat8-CD147Ex and to Mr. Phawin Charoensook for technical assistance. We also acknowledge Dr. Ian Thomas and Mr. Brian William Habbard for critical reading of this manuscript.

Author information

Correspondence to Chatchai Tayapiwatana.

Additional information

This work was supported by the Thailand Research Fund and the National Center for Genetic Engineering and Biotechnology. R.J.T. thanks the Canadian Institutes of Health Research for funding support.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Thammawong, P., Kasinrerk, W., Turner, R.J. et al. Twin-arginine signal peptide attributes effective display of CD147 to filamentous phage. Appl Microbiol Biotechnol 69, 697–703 (2006). https://doi.org/10.1007/s00253-005-0242-0

Download citation

Keywords

  • Phage Display
  • Sandwich ELISA
  • Phage Particle
  • Filamentous Phage
  • Phage Display Technique