SEMINAR

Structural Basis for Antibody Recognition of an Intrinsically Disordered Antigen

Abstract
Intrinsically disordered proteins are highly abundant in Plasmodium and related pathogenic genera.1 Merozoite surface protein 2 (MSP2) is an intrinsically disordered, GPI-anchored membrane antigen of the malaria parasite Plasmodium falciparum,2 which elicits a protective immune response in humans. We have undertaken a detailed analysis of the conformational dynamics of the two major allelic forms of MSP2 (3D7 and FC27) using NMR spectroscopy. Chemical shifts and NMR relaxation data indicate that the conformational and dynamic properties of the N- and C-terminal conserved regions in the two forms of MSP2 are essentially identical, although significant variation exists between and within the central variable regions.3 We observe a strong relationship between the conformational dynamics and the antigenicity of MSP2, as assessed with antisera to recombinant MSP2. Regions of increased conformational order in MSP2, including those in the conserved regions, are more strongly antigenic, while the most flexible regions are minimally antigenic.

Antibodies are generated to the conserved N- and C-terminal regions of soluble, recombinant MSP2, but many of these react poorly with the native antigen on the parasite surface. We have shown4 that recognition of a conserved N-terminal epitope by mAb 6D8 is incompatible with the membrane-bound conformation of that region,5 suggesting a mechanism by which native MSP2 on the parasite escapes antibody recognition. Furthermore, crystal structures and NMR spectroscopy identify transient (‘fuzzy’), strain-specific interactions between the 6D8 antibody and regions of MSP2 beyond the conserved epitope. These interactions account for the differential affinity of 6D8 for the two allelic families of MSP2, even though 6D8 binds to a fully conserved epitope.

References
1 Feng, Z. P. et al. Mol Biochem Parasitol 150, 256-267 (2006).
2 Zhang, X. et al. J Mol Biol 379, 105-121 (2008).
3 MacRaild, C. A. et al. PLoS One 10, e0119899 (2015).
4 Morales, R. A. V. et al. Sci Rep 5, 10103 (2015).
5 MacRaild, C. A. et al. Biochim Biophys Acta 1818, 2572-2578 (2012).

BIOGRAPHY

2010-present: Professor, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville.
2004-present: NHMRC Principal Research Fellow.
2001-2010: Laboratory Head and Deputy Division Head, Structural Biology Division, The Walter and Eliza Hall Institute of Medical Research.
1992-2000: Head, NMR Laboratory and Assistant Director, Biomolecular Research Institute, Melbourne.
1981-1992: Lecturer, Senior Lecturer, Associate Professor, School of Biochemistry, University of NSW, Sydney.
1974: PhD, Australian National University, Canberra.
1971: BSc (Hons), University of Melbourne, Melbourne.

Research Fields and Interests:
Professor Ray Norton holds a personal chair at the Monash Institute of Pharmaceutical Sciences, Parkville. His lab employs a range of biophysical approaches, including NMR, SPR, ITC and X-ray crystallography, in studies of peptide and protein toxins and malaria. One of the peptide toxins he works with is about to enter phase II clinical trial for autoimmune diseases. This range of biophysical methods is also applied in combined structure-based and fragment-based drug discovery programs against protein targets in the areas of infectious and parasitic diseases, graft rejection and metabolic diseases. He has published over 310 articles and is an inventor on 10 patents.

Selected Recent Publications:
Rashid MH, Huq R, Tanner MR, Chhabra S, Khoo KK, Estrada R, Dhawan V, Chauhan S, Pennington MW, Beeton C, Kuyucak S & Norton RS (2014) A potent and Kv1.3-selective analogue of the scorpion toxin HsTX1 as a potential therapeutic for autoimmune diseases. Scientific Reports 4, 4509 (9 pages).
Yap BK, Leung EWW, Yagi H, Chhabra S, Chalmers DK, Nicholson SE, Thompson PE & Norton RS (2014) A potent cyclic peptide targeting SPSB2 protein as a potential anti-infective agent. Journal of Medicinal Chemistry 57, 7006-7015.
Chhabra S, Chang SC, Nguyen HM, Huq R, Tanner MR, Londono LM, Estrada R, Dhawan V, Chauhan S, Upadhyay SK, Gindin M, Hotez PJ, Valenzuela JG, Mohanty B, Swarbrick JD, Wulff H, Iadonato SP, Gutman GA, Beeton C, Pennington MW, Norton RS* & Chandy KG* (2014) Kv1.3 channel-blocking immunomodulatory peptides from parasitic worms: implications for autoimmune diseases. * joint corresponding authors FASEB Journal 28, 3952-3964.
Safavi-Hemami H, Gajewiak J, Karanth S, Robinson SD, Ueberheide B, Douglass A, Schlegel A, Imperial J, Watkins M, Bandyopadhyay PK, Yandell M, Li Q, Purcell AW, Norton RS, Ellgaard L & Olivera BM (2015) A specialized insulin is used for chemical warfare by fish-hunting cone snails. Proceeding of the National Academy of Sciences USA, 112, 1743-1748.
Morales RAV, MacRaild CA, Seow J, Krishnarjuna B, Drinkwater N, Rouet R, Anders RF, Christ D, McGowan S & Norton RS (2015) Structural basis for epitope masking and strain specificity of a conserved epitope in an intrinsically disordered malaria vaccine candidate. Scientific Reports 5, 10103 (10 pages).