The characterisation of the peanut agglutinin an evolved plant lectin, with improved specificity to the Thompson Freidenriech antigen

Peanut agglutinin (PNA), a carbohydrate binding protein, is able to recognise and bind a number of distinct carbohydrate structures that have been implicated in a number of disease pathologies in humans. In vitro studies of PNA have previously been shown to have some specificity for the Thomson Freidenriech antigen (T-antigen), found on malignant human cells, and this specificity has made PNA an important target for protein engineering experiments aimed at improving its specificity and affinity. A number of tumour cells are characterised by altered states and patterns of glycosylation on cell surfaces and suitably engineered lectins may be able to recognise tumour specific carbohydrate structures. This study was aimed at carrying out the biophysical characterisation of a set of PNA mutants which showed apparent improvement in specificity for the T-Antigen. Previous studies have aimed to engineer this lectin in order to direct its recognition properties towards the T-antigen and away from lactose, the preliminary binding affinities of these mutants being determined using Surface Plasmon Resonance (SPR). Here a set of PNA mutants were characterised, proteins expressed and purified to determine binding activities to the T-antigen, N-Acetyl-Dlactosamine (LacNAc) and lactose through the use of Protein Micro Array technology as well as Enzyme linked immunosorbant assays (ELISA).

Grantor dc:publisher.institution

Division of Medical Biochemistry

Year dc:date.issued

2013

Author dc:creator

• Lagardien, Zaida

Advisor dc:contributor.advisor

• Blackburn, Jonathan

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/11427/3136

OAI identifier oai:identifier

oai:open.uct.ac.za:11427/3136