TEV NIa protease as a tool for in vitro and in vivo proteolysis

Maltose transporter
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Gene expression in
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TEV NIa protease
as a tool

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© 2001 Webdesign by
Tilman Maier og Lars Jessen
1. Introducing a TEV protease recognition site
2. Expression vectors
3. Purification and storage of TEV protease
4. Proteolysis in vitro
5. Proteolysis of whole cells
6. Proteolysis in vivo
7. References
8. Links

7. References

  • Characterisation of TEV protease and its recognition sites

    Carrington, J. C., and W. G. Dougherty. 1988. A viral cleavage site cassette: identification of amino acid sequences required for tobacco etch virus polyprotein processing. Proc. Natl. Acad. Sci. USA 85: 3391-3395.

    Dougherty, W. G., S. M. Cary, and T. D. Parks. 1989. Molecular genetic analysis of a plant virus polyprotein cleavage site: a model. Virology 171: 356-364.

    Parks, T. D., H. A. Smith, and W. G. Dougherty. 1992. Cleavage profiles of tobacco etch virus (TEV)-derived substrates mediated by precursor and processed forms of the TEV NIa proteinase. J. Gen. Virol. 73:149-155

    Dougherty, W., and B. Semler. 1993. Expression of Virus-Encoded Proteinases: Functional and Structural Similarities with Cellular Enzymes. Microbiol. Rev. 57: 781-822.

    Mohanty AK, Simmons CR, Wiener MC. 2003 Inhibition of tobacco etch virus protease activity by detergents. Protein Expr Purif.27:109-14. entrez/medline

    Tozser J, Tropea JE, Cherry S, Bagossi P, Copeland TD, Wlodawer A, Waugh DS. 2005 Comparison of the substrate specificity of two potyvirus proteases. FEBS J.272:514-23. entrez/medline

  • in vivo proteolysis

    Yang X, Gregan J, Lindner K, Young H, Kearsey SE. 2005 Nuclear distribution and chromatin association of DNA polymerase alpha-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast. BMC Mol Biol. 6:13 biomed central

    Henrichs T, Mikhaleva N, Conz C, Deuerling E, Boyd D, Zelazny A, Bibi E, Ban N, Ehrmann M. 2005 Target-directed proteolysis at the ribosome. Proc Natl Acad Sci U S A. 102:4246-51. entrez/medline

    Herskovits, A.A., Seluanov, A., Rajsbaum, R., ten Hagen-Jongman, C.M., Henrichs, T., Bochkareva, E.S., Phillips, G.J., Probst, F.J., Nakae, T., Ehrmann, M., Luirink, J., and Bibi, E. 2001. Evidence for coupling of membrane- targeting and function of the signal recognition particle-receptor FtsY. EMBO Reports, 2:1040-1046 entrez/medline

    Ehrmann, M., Bolek, P., Mondigler, M., Boyd, D., and R. Lange 1997. TnTIN and TnTAP: mini-transposons for site specific proteolysis in vivo. Proc. Natl. Acad. Sci. USA 94:13111-13115 entrez/medline

    Mondigler M. and M. Ehrmann. 1996. Site specific proteolsis of the E. coli SecA Protein in vivo. J. Bacteriol. 178: 2986-2988 entrez/medline

    Kapust RB, Waugh DS. 2000. Controlled intracellular processing of fusion proteins by TEV protease. Protein. Expr. Purif. 2000 19:312-318

  • Expression in yeast

    Yang X, Gregan J, Lindner K, Young H, Kearsey SE. 2005 Nuclear distribution and chromatin association of DNA polymerase alpha-primase is affected by TEV protease cleavage of Cdc23 (Mcm10) in fission yeast. BMC Mol Biol. 6:13 biomed central

    Faber, K.N., Kram, AM., Ehrmann, M., and Veenhuis, M. 2001. The use of the Tobacco Etch Virus (TEV-) protease as a tool to determine the topology of peroxisomal membrane proteins in vivo. J. Biol. Chem. 276: 36501-36507 entrez/medline

    Uhlmann F, Wernic D, Poupart MA, Koonin EV, Nasmyth K. 2000. Cleavage of cohesin by the CD clan protease separin triggers anaphase in yeast. Cell 103:375-386

    Smith, T. A. & Kohorn, B. D. 1991. Direct selection for sequences encoding proteases of known specificity. Proc. Natl. Acad. Sci. USA. 88: 5159-5162.

  • Cleavage of hybrid proteins

    Parks, T. D., Leuther, K. K., Howard, E. D., Johnston, S. A. & Dougherty, W. G. 1994. Release of proteins and peptides from fusion proteins using a recombinant plant virus proteinase. Anal. Biochem. 216: 413-417.

  • Production of TEV protease

    Kapust RB, Routzahn KM, Waugh DS. 2002. Processive Degradation of Nascent Polypeptides, Triggered by Tandem AGA Codons, Limits the Accumulation of Recombinant Tobacco Etch Virus Protease in Escherichia coli BL21(DE3). Protein. Expr. Purif. 24:61-70

    Kapust RB, Tozser J, Fox JD, Anderson DE, Cherry S, Copeland TD, Waugh DS. 2001. Tobacco etch virus protease: mechanism of autolysis and rational design of stable mutants with wild-type catalytic proficiency. Protein. Eng. 14:993-1000

    Lucast, LJ, Batey, RT, and Doudna, JA. 2001 Large-scale purification of a stable form of recombinant tobacco etch virus protease. Biotechniques 30: 544-546

  • Expression vectors containing TEV protease sites

    Melcher K. 2000. A modular set of prokaryotic and eukaryotic expression vectors. Anal Biochem 277:109-120
    Erratum in: Anal Biochem 2000. 282:266

  • TEV protease used to study protein localisation

    Gao XD and Dean N. 2000. Distinct protein domains of the yeast Golgi GDP-mannose transporter mediate oligomer assembly and export from the endoplasmic reticulum. J. Biol. Chem. 275:17718-17727

  • TEV protease used to study bacterial outer membrane proteins

    Guedin, S., Willery, E., Tommassen, J., Fort, E., Drobecq, H., Locht, ., and Jacob-Dubuisson, F. 2000. Novel Topological Features of FhaC, the Outer Membrane Transporter Involved in the Secretion of the Bordetella pertussis Filamentous Hemagglutinin. J. Biol. Chem. 275: 30202-30210

    Guilvout, I., Hardie, K. R., Sauvonnet, N., and Pugsley, A.P. 1999. Genetic Dissection of the Outer Membrane Secretin PulD: Are There Distinct Domains for Multimerization and Secretion Specificity? J. Bacteriol. 181:7212-7220.

  • Other applications

    Marcos, J., and R. Beachy. 1994. In vitro characterization of a cassette to accumulate multiple proteins through synthesis of a self-processing polypeptide. Plant Mol. Biol. 24: 495-503.