Plant plasmalemma structure: an immunological approach

Norman, Paul Michael (1986). Plant plasmalemma structure: an immunological approach. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000f81d

Abstract

Clonal hybridomas were generated which secreted monoclonal antibodies reactive with a crude membrane preparation from Nicotiana glutinosa suspension culture cells. Antibody secretion was assessed by a radioimmunoassay using such membranes as substrate. A number of the monoclonal antibodies recognised epitopes expressed on the external face of the plasmalemma of N. glutinosa suspension culture derived protoplasts, as assessed by immunofluorescence microscopy and flow cytometry. A second, non-overlapping set of antibodies recognised epitopes on the exterior face of the cell wall of intact cells, whilst a third group showed neither reactivity, and was postulated to recognise epitopes expressed within the cell.

Western blotting and immunoprecipitation analyses identified antibodies recognising a number of proteins including several plasmalemma glycoproteins. The recognised epitopes were periodate sensitive, and so probably in carbohydrate moieties. Immunoaffinity chromatography of a detergent extract of plant cells allowed purification of a plasmalemma glycoprotein. This was subjected to amino acid analysis, and used to raise polyclonal antisera and further monoclonal antibodies. Deglycosylation of a partially purified detergent extract of N. glutinosa cells suggested that this plasmalemma glycoprotein consists of a 50 kd molecular weight core protein, which is extensively and heterogeneously glycosylated, raising its apparent molecular weight to 130-230 kd.

The plasmalemma glycoprotein was used as an antigenic marker for plasmalemma derived vesicles resolved on sucrose density gradients, and for heterokaryons in protoplast fusions. Similarities between the plasmalemma glycoprotein and arabinogalactan proteins are discussed.

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