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Cloning and expression of truncated form of tissue plasminogen activator in Leishmania tarentolae

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Abstract

An expression cassette containing kringle 2 and serine protease domains (K2S), tissue plasminogen activator (tPA), together with a signal sequence derived from Leishmania tarentolae and two fragments of the small subunit ribosomal RNA locus, was introduced into L. tarentolae. The transfected cells produced recombinant K2S (rK2S) protein extracellularly with serine protease activity. Expression and enzyme activity of rK2S in the supernatant was 930 i.u./ml. The specific activity of purified rK2S was 7.4 U/mg of protein. Replacement of the human signal sequence tPA with the signal sequence derived from Leishmania increased the secretion of recombinant protein up to 30 times.

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References

  • Baruah DB, Dash RN, Chaudhari MR et al (2006) Plasminogen activators: a comparison. Vascul Pharmacol 44:1–9

    Article  CAS  PubMed  Google Scholar 

  • Breitling R, Klingner S, Callewaert N et al (2002) Non-pathogenic trypanosomatid protozoa as a platform for protein research and production. Protein Expr Purif 25:209–218

    Article  CAS  PubMed  Google Scholar 

  • Clayton CE (2002) Life without transcriptional control? From fly to man and back again. EMBO J 21:1881–1888

    Article  CAS  PubMed  Google Scholar 

  • Ellis K, Brener S (2004) New fibrinolytic agents for MI: as effective as current agents, but easier to administer. Clevel Clin J Med 71(20):23–25 29–30 passim

    Google Scholar 

  • Georgiou G, Valax P (1996) Expression of correctly folded proteins in Escherichia coli. Curr Opin Biotechnol 7:190–197

    Article  CAS  PubMed  Google Scholar 

  • Heussen C, Dowdle EB (1980) Electrophoretic analysis of plasminogen activators in polyacrylamide gels containing sodium dodecyl sulfate and copolymerized substrates. Anal Biochem 102:196–202

    Article  CAS  PubMed  Google Scholar 

  • Hockney RC (1994) Recent developments in heterologous protein production in Escherichia coli. Trends Biotechnol 12:456–463

    Article  CAS  PubMed  Google Scholar 

  • Kohnert U, Rudolph R, Verheijen JH et al (1992) Biochemical properties of the kringle 2 and protease domains are maintained in the refolded t-PA deletion variant BM 06.022. Protein Eng 5:93–100

    Article  CAS  PubMed  Google Scholar 

  • La Flamme AC, Buckner FS, Swindle J et al (1995) Expression of mammalian cytokines by Trypanosoma cruzi indicates unique signal sequence requirements and processing. Mol Biochem Parasitol 75:25–31

    Article  CAS  PubMed  Google Scholar 

  • Manosroi J, Tayapiwatana C, Götz F et al (2001) Secretion of active recombinant human tissue plasminogen activator derivatives in Escherichia coli. Appl Environ Microbiol 67:2657–2664

    Article  CAS  PubMed  Google Scholar 

  • Mattes R (2001) The production of improved tissue-type plasminogen activator in Escherichia coli. Semin Thromb Hemost 27:325–336

    Article  CAS  PubMed  Google Scholar 

  • Misslitz A, Mottram JC, Overath P et al (2000) Targeted integration into a rRNA locus results in uniform and high level expression of transgenes in Leishmania amastigotes. Mol Biochem Parasitol 107:251–261

    Article  CAS  PubMed  Google Scholar 

  • Nordt TK, Bode C (2003) Thrombolysis: newer thrombolytic agents and their role in clinical medicine. Heart 89:1358–1362

    Article  CAS  PubMed  Google Scholar 

  • Orlando TC, Mayer MG, Campbell DA et al (2007) RNA polymerase I promoter and splice acceptor site recognition affect gene expression in non-pathogenic Leishmania species. Mem Inst Oswaldo Cruz 102:891–894

    Article  CAS  PubMed  Google Scholar 

  • Pennica D, Holmes WE, Kohr WJ et al (1983) Cloning and expression of human tissue-type plasminogen activator cDNA in E. coli. Nature 301:214–221

    Article  CAS  PubMed  Google Scholar 

  • Rouf SA, Moo-Young M, Chisti Y (1996) Tissue-type plasminogen activator: characteristics, applications and production technology. Biotechnol Adv 14:239–266

    Article  CAS  PubMed  Google Scholar 

  • Soleimani M, Mahboudi F, Davoudi N et al (2007) Expression of human tissue plasminogen activator in the trypanosomatid protozoan Leishmania arentolae. Biotechnol Appl Biochem 48:55–61

    Article  CAS  PubMed  Google Scholar 

  • Tobin JF, Reiner SL, Hatam F et al (1993) Transfected Leishmania expressing biologically active IFN-gamma. J Immunol 150:5059–5069

    CAS  PubMed  Google Scholar 

Download references

Acknowledgment

This work was supported by a grant from the Pasteur Institute of Iran.

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Authors and Affiliations

  1. Department of Microbiology, Qom Branch, Islamic Azad University, 15 Khordad BLVD, Qom, Iran

    Razieh Nazari

  2. Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran

    Noushin Davoudi

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  1. Razieh Nazari
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  2. Noushin Davoudi
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Correspondence to Razieh Nazari.

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Nazari, R., Davoudi, N. Cloning and expression of truncated form of tissue plasminogen activator in Leishmania tarentolae . Biotechnol Lett 33, 503–508 (2011). https://doi.org/10.1007/s10529-010-0470-y

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  • DOI: https://doi.org/10.1007/s10529-010-0470-y

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