Abstract
Proteinase inhibitors (PIs) of the potato type II family have been identified in a number of solanaceous species. Most family members have two PI domains which are specific for either chymotrypsin or trypsin. More recently family members have been described with three or six repeated PI domains. Here we describe a novel four-domain family member produced in the stigmas and leaves of the ornamental tobacco, Nicotiana alata, which has high sequence identity with a six-domain member from the same species. Both proteins are produced as precursors that enter the secretory pathway and are subsequently processed into a series of 6 kDa PIs. The four- and six-domain precursor proteins were isolated from immature stigmas and characterised by mass spectrometry which revealed that both proteins had been trimmed at the N-terminus, at a position corresponding to the predicted signal peptide cleavage site. Furthermore, no post-translational modifications were apparent.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Atkinson, A.H., Heath, R.L., Simpson, R.J., Clarke, A.E. and Anderson, M.A. 1993. Proteinase inhibitors in Nicotiana alata stigmas are derived from a precursor protein which is processed into five homologous inhibitors. Plant Cell 5: 203–213.
Baladin, R., van der Does, C., Albert, J.M.B., Bol, J.F. and Linthorst, H.J.M. 1995. Structure and induction pattern of a novel proteinase inhibitor class II gene of tobacco. Plant Mol. Biol. 27: 1197–1204.
Bryant, J., Green, R.T., Gurusaddiaiah, T. and Ryan, C.A. 1976. Proteinase inhibitor II from potatoes: isolation and characterization of its promoter components. Biochemistry 15: 3418–3424.
Graham, J.S., Pearce, G., Merryweather, F., Titani, K., Ericsson, L.H. and Ryan, C.A. 1985. Wound-induced proteinase inhibitors from tomato leaves. II. The cDNA-deduced primary structure of pre-inhibitor II. J. Biol. Chem. 260: 6561–6564.
Hass, G.M., Hermodson, M.A., Ryan, C.A. and Gentry, L. 1982. Primary structures of two low molecular weight proteinase inhibitors from potatoes. Biochemistry 21: 752–756.
Heath, R.L., Barton, P.A., Reid, G.E.M., Lim, G. and Anderson, M.A. 1995. Characterisation of the protease processing sites in a multidomain proteinase inhibitor precursor from Nicotiana alata. Eur. J. Biochem. 230: 250–257.
Heath, R.L., McDonald, G., Christeller, J.T., Lee, M., Bateman, K., West, J., van Heeswijk, R. and Anderson, M.A. 1997. Proteinase inhibitors from Nicotiana alata enhance resistance to insect pests. J. Insect Physiol. 43: 833–842.
Keil, M., Sanchez-Serrano, J., Schell, J. and Willmitzer, L. 1986. Primary structure of a proteinase inhibitor II gene from potato. Nucl. Acids Res. 14: 5641–5650.
Lee, M.C.S., Scanlon, M.J., Craik, D.J. and Anderson, M.A. 1999. A novel two-chain proteinase inhibitor generated by circularization of a multidomain precursor protein. Nature Struct. Biol. 6: 526–530.
Miller, E.A., Lee, M.C.S. and Anderson, M.A. 1999. Identification and characterization of a prevacuolar compartment in stigmas of Nicotiana alata. Plant Cell 11: 1499–1508.
Nielsen, K.J., Hill, J.M., Anderson, M.A. and Craik, D.J. 1996. Synthesis and structure determination by NMR of a putative vacuolar targeting peptide and model of a proteinase inhibitor from Nicotiana alata. Biochemistry 35: 369–378.
Pearce, G., Johnson, S. and Ryan, C.A. 1993. Purification and characterization from tobacco (Nicotiana tabacum) leaves of six small, wound-inducible, proteinase isoinhibitors of the potato inhibitor II family. Plant Physiol. 102: 639–644.
Plunkett, G., Senear, D.F., Zuroske, G. and Ryan, C.A. 1982. Proteinase inhibitors I and II from leaves of wounded tomato plants: purification and properties. Arch. Biochem. Biophys. 213: 463–472.
Richardson, M. 1979. The complete amino acid sequence and the trypsin reactive (inhibitory) site of the major proteinase inhibitor from the fruits of aubergine (Solanum melongena L.). FEBS Lett. 104: 322–326.
Sanchez-Serrano, J., Schmidt, R., Schell, J. and Willmitzer, L. 1986. Nucleotide sequence of proteinase inhibitor II encoding cDNA of potato (Solanum tuberosum) and its mode of expression. Mol. Gen. Genet. 203: 15–20.
Scanlon, M.J., Lee, M.C.S., Anderson, M.A. and Craik, D.J. 1999. Structure of a putative ancestral protein encoded by a single sequence repeat from a multidomain proteinase inhibitor gene from Nicotiana alata. Structure 7: 793–802.
Schultz, C.J., Hauser, K., Lind, J.L., Atkinson, A.H., Pu, Z.Y., Anderson, M.A. and Clarke, A.E. 1997. Molecular characterisation of a cDNA sequence encoding the backbone of a style-specific 120 kDa glycoprotein which has features of both extensins and arabinogalactan proteins. Plant Mol. Biol. 35: 833–845.
Taylor, B.H., Young, R.J. and Scheuring, C.F. 1993. Induction of a proteinase inhibitor II-class gene by auxin in tomato roots. Plant Mol. Biol. 23: 1005–1014.
Thornberg, R.W., An, G., Cleveland, T.E., Johnson, R. and Ryan, C.A. 1987. Wound inducible expression of a potato inhibitor II-chloramphenicol acetyltransferase gene fusion in transgenic tobacco plants. Proc. Natl. Acad. Sci. USA 84: 744–748.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Miller, E.A., Lee, M.C., Atkinson, A.H. et al. Identification of a novel four-domain member of the proteinase inhibitor II family from the stigmas of Nicotiana alata. Plant Mol Biol 42, 329–333 (2000). https://doi.org/10.1023/A:1006305429013
Issue Date:
DOI: https://doi.org/10.1023/A:1006305429013