PEPTIDE DRUG DESIGN AND MEMBRANE NANOTECHNOLOGY

Professor Mibel Aguilar

Email: mibel.aguilar@monash.edu

Our group focuses on peptide-based drug design and biomembrane nanotechnology. In collaboration with Prof Patrick Perlmutter (Chemistry), we are developing novel compounds that allow us to exploit the potential of peptides as drugs. We are currently applying our technology to the development of cancer vaccines (with Assoc Prof Tony Purcell, Uni Melb), and new compounds for treatments of cardiovascular disease (with Prof Rob Widdop) and HIV (with Assoc Prof Gilda Tachedjian, Burnet Institute and Dr Miranda Shehu-Xhilaga). Our membrane nanotechnology projects involve the development of new methods for membrane protein purification and analysis with application to Alzheimer's (with Prof David Small, Menzies Research Institute), G protein-coupled receptor function with Prof Wally Thomas (University of Qld), antimicrobial peptide function (with Prof Frances Separovic (Uni Melb) and new biosensor devices (with Farfield Scientific). We also collaborate with Dr Mary Ann Augustin and Dr Li Day (CSIRO, Food & Nutritional Sciences) on applications of protein nanotechnology to improving food formulations.

The long-term aim of these studies is to increase our understanding of the molecular basis of peptide and protein function and allow the rational design of peptide and protein based therapeutics.

MEMBRANE NANOTECHNOLOGY

The interactions between peptides and lipids are of fundamental importance in the functioning of numerous membrane-mediated biochemical processes including antimicrobial peptide action, hormone-receptor interactions, drug bioavailability across the blood-brain barrier and viral fusion processes. Indeed, a major target of modern biotechnology is the design of new potent pharmaceutical agents whose biological action is dependent on the binding of peptides with lipid-bilayers.

Model of the interaction of different peptides & proteins with the plasma membrane.

Antimicrobial Peptide Action

Antimicrobial peptides are being increasingly recognised as potential candidate antibacterial drugs in the face of the rapidly emerging bacterial resistance to conventional antibiotics in recent years. However, a precise understanding of the relationship between antimicrobial peptide structure and their cytolytic function in a range of organisms is still lacking. This is a result of the complex nature of the interactions of antimicrobial peptides with the cell membrane, the mechanism of which can vary considerably between different classes of antimicrobial peptides. Antimicrobial peptide action is mediated by a direct interaction with cell membranes and a common feature of these interactions is the induction of cationic amphipathic secondary structure following binding of the peptides to the membrane surface. Since selective binding to different phospholipids is central to the design of non-hemolytic antimicrobial peptides, the affinity of the peptide for the membrane surface is a critical factor in the cell-lytic process. We have developed a sensitive method based on surface plasmon resonance (SPR), which allows the real-time measurement of peptide binding to phospholipid membranes. The project area focuses on characterising the molecular basis of antimicrobial peptide action to assist the direct design of more potent and selective antimicrobial peptides as leads from new therapeutics for the treatment of bacterial infections.

Adsorption	Adsorption	Pore formation

Publications:

• Lee TH, and Aguilar MI. ‘Dual Polarisation Interferometry: An Optical Biosensor Which Allows New Insights into Peptide-Induced Changes in Biomembrane Structure', Aust J Chem., (2011), in press.
• Hall K, Lee TH and Aguilar MI, ‘The Role of Electrostatic interactions in the Membrane Binding of Melittin', J Mol Recog, 24 (2011) 108-118.
• Hirst D, Lee TH, Swann MJ, Unabia S, Park YK, Hahm, KS and Aguilar MI, ‘Effect of acyl chain structure and bilayer phase state on the binding and penetration of a supported lipid bilayer by HPA3', Eur Biophys J, 40 (2011) 503-514.
• Lee TH, Hall K, Swann MJ, Popplewell JF, Park YK, Hahm, KS and Aguilar MI, ‘The Membrane Insertion of Helical Antimicrobial Peptides from the N-Terminus of Helicobacter pylori Ribosomal Protein L1', Biochim Biophys Acta (Biomembranes), 1798 (2010) 544-557.
• Lee TH, Heng C, Swann MJ, Gehman JD, Separovic F and Aguilar MI, ‘Real time quantitative analysis of lipid disordering by aurein 1.2 during membrane adsorption, destabilisation and lysis', Biochim Biophys Acta (Biomembranes), 1798 (2010) 1977-1986.
• Henriques ST, Pattenden LK, Castanho MARB, Aguilar MI, ‘Fast membrane association is a crucial factor in the pep-1 translocation mechanism: A kinetic study followed by surface plasmon resonance', Biopolymers-Pept Sci, 94 (2010) 314-322.
• Hall K and Aguilar MI, ‘Membrane interaction of antimicrobial β-peptides and the role of amphipathicity', Biopolymers-Pept Sci., 92 (2009) 554-564.
• Veiga AS, Pattenden LK, Fletcher JM, Castanho MARB and Aguilar MI, ‘Real-time Biosensor Interactions of HIV-1 Antibodies 2F5 and 4E10 with a gp41 Epitope Prebound to Host and Viral Membrane Mimetics'. Chem Biochem, 10 (2009) 1032-1044. (IF 3.446, Cited 6).
• Mechler A, Praporski S, Piantavigna S, Heaton SM, Hall KN, Aguilar MI and Martin LL^, ‘Pseudo-physiological phospholipid bilayers: structure, homogeneity and deposition
  characteristics on carboxylic acid terminated self-assembled monolayers', Biomaterials, 30 (2009) 682-689.
• Henriques ST, Pattenden LK, Aguilar MI, Castanho MARB, ‘The toxicity of prion protein fragment PrP(106-126) is Not Mediated by Membrane Permeabilization as shown by a M112W substitution'. Biochemistry, 48 (2009) 4198-4208
• Gehman JD, Luck F. Hall K, Lee, TH, Boland MP, Pukala TL, Bowie JH, Aguilar MI, Separovic F. ‘Effect of antimicrobial peptides from Australian tree frogs on anionic phospholipid membranes'. Biochemistry 47 (2008) 8557-8567.
• Kamimori H, Hall K, Craik, DJ, Aguilar MI, ‘Studies on the Membrane Interactions of the Cyclotides Kalata B1 and Kalata B6 on Model Membrane Systems by Surface Plasmon Resonance. Anal Biochem, 337 (2004) 149-153.
• Hall, K, Mozsolits, H, Aguilar MI, ‘Surface Plasmon Resonance Analysis of Antimicrobial Peptide-Membrane Interactions: Affinity and Mechansim of Action', Lett. In Pep. Sci., 10 (2004), 475-485.
• Mozsolits H, Lee T.H, Clayton A, Sawyer, W.H. and Aguilar M.I., `The membrane binding properties of a Class A amphipathic peptide', Eur Biophys J, 270 (2003), 4282-93.
• Jin Y., Mozsolits H., Hammer J., Zmuda E., Zhu J., Zhang Y., Aguilar M.I. and Blazyk, J., `Influence of tryptophan on lipid-binding of linear amphipathic cationic antimicrobial peptides', Biochemistry 42 (2003) 9395-405.
• Mozsolits H and Aguilar M I, `Surface Plasmon Resonance Spectroscopy: An Emerging Tool for the Study of Peptide-Membrane Interactions', Biopolymers - Peptide Science, 66 (2002) 3-18.

• Mozsolits H, Wirth H-J, Werkmeister J and Aguilar M.I., `Analysis of antimicrobial peptide interactions with hybrid bilayer membrane systems using surface plasmon resonance', Biochim Biophys Acta 1512 (2001) 64-76.

G Protein-Coupled Receptor (GPCR) Regulation

Seven transmembrane-spanning receptors that couple to heterotrimeric G proteins represent by far the largest receptor superfamily in our genome, mediating functions across the spectrum of physiology.  Elucidating the mechanisms that activate/deactivate these G protein-coupled receptors (GPCRs) is fundamental to expanding our understanding of these receptors. Moreover, many therapeutics aim to target GPCRs and hence developing an accurate picture of their function is a continuing focus of the pharmaceutical industry. The participation of the plasma membrane in GPCR signaling and regulation is emerging as a key feature of GPCR structure and function and we aim to characterise the molecular details of this synergy.

In collaboration with Prof Wally Thomas (University of Queensland) we are exploring the role of membranes in the study of membrane mediated receptor activation using the G-protein coupled angiotensin (AT1) receptor as a model system. GPCRs are integral membrane proteins with a structure consisting of an extracellular amino-terminus, seven transmembrane-spanning b-helices connected by alternating extracellular and intracellular loops, and a cytoplasmic carboxyl-terminus. The AT1 receptor is a 359 amino acid GPCR that mediates the important cardiovascular and homeostatic actions of the peptide hormone, angiotensin II and it has been shown that receptor expression, affinity, signaling and trafficking is crucially dependent upon the membrane interactions.

Publications:

• Du AT, Onan D, Dinh DT, Lew MJ, Ziogas J, Aguilar MI, Pattenden LK and Thomas WG, ‘Ligand supported purification of the urotensin-II receptor', Mol Pharmacol, 78 (2010) 639-647.
• Huynh J, Thomas, WG, Aguilar MI, Pattenden LK, ‘The Type 1 Angiotensin Receptor: the Role of Helix 8 in G Protein-Coupled Receptor Structure and Function', Mol Cell Endo, 302 (2009) 118-127.
• Kamimori H, Unabia, S, Thomas, WG, Aguilar MI, Evaluation of the membrane-binding properties of the Proximal Region of the Angiotensin II Receptor (AT_1A) Carboxyl Terminus by Surface Plasmon Resonance', Anal Sci., 21 (2005) 171-4.
• Mozsolits, H., Unabia, S., Ahmad, A., Morton, C.J., Thomas, W.G., Aguilar, M.-I. Electrostatic and hydrophobic forces tether the proximal region of the angiotensin II receptor (AT_1A) carboxyl-terminus to the cell membrane. Biochemistry, 41 (2002) 7830-7840.
• Mozsolits, H., Thomas, W.G., Aguilar, M.-I. ‘Surface plasmon resonance spectroscopy in the study of membrane-mediated cell signalling', J Pept Sci, , 66 (2002) 3-18.

Role of Membrane Binding in Neurodegenerative Diseases

Understanding the mechanism by which accumulation of the beta-amyloid protein (Ab) in the brain contributes to the onset of dementia is one of the main unsolved problems in the field of Alzheimer's disease (AD) research. In collaboration with Prof David Small, we have recently identified a novel mechanism by which Ab exerts its toxic effects via direct binding with the cell membrane, an effect which is mediated by the presence of cholesterol, a known risk factor in AD.

In particular, our studies provide a molecular snapshot of Ab formation and aggregation and suggest that Ab binding to membranes is a potential therapeutic target for the treatment of Alzheimer's disease. We are now applying our membrane biosensor techniques together with atomic force microscopy, to studying the role of membrane-binding in the cellular toxicity associated with amyloid formation in Alzheimer's disease. We are also extending these studies to a number of other neurodegenerative diseases associated with protein misfolding including transthyretin which has been demonstrated to be the predominant component of the amyloid fibrils in familial amyloidotic polyneuropathy.

Publications:

• Klaver D, Hung AC, Gasperini R, Foa L, Aguilar MI and Small DH, ‘Glycosaminoglycan-BACE1 interactions as a target for drug development in Alzheimer's disease' Neurodegenerative Diseases, 7 (2010) 187-189.
• Henriques ST, Pattenden LK, Aguilar MI, Castanho MARB, ‘Neurotoxicity of PrP(106-126) revisited. ‘PrP(106-126) does not interact with membranes under physiological conditions'. Biophys J, 95 (2008) 1877-89
• Hou X, Mechler A, Martin LL, Aguilar MI, Small DH. ‘Cholesterol and anionic lipids increase the binding of amyloidogenic transthyretin to lipid membranes'. Biochim Biophys Acta, 1178 (2008) 198-205.
• Petratos S, Li QX, George AJ, Hou X, Kerr M, Unabia S, Hatzinisiriou I, Maksel D, Aguilar MI, Small DH. ‘The b-amyloid protein of Alzheimer's disease increases neuronal CRMP-2 phosphorylation by a Rho-GTP mechanism', Brain, 131 (2008) 90-108.
• Hou X, Parkington, HC, Coleman, HA, Mechler, A, Martin, LL, Aguilar MI, & Small, DH, ^"Transthyretin oligomers induce calcium influx via voltage-gated calcium channels", J Neurochem, 100, (2007) 446-57.
• Small, DH, Maksel, D, Ng, J, Kerr, ML, Hou, X, Chu, C, Mehrani, H, Unabia, S, Azari, MF, Loiacono, R, Aguilar MI and Chebib, M. "The b-amyloid protein of Alzheimer's disease binds to membrane lipids but does not bind to the a-7 nicotinic acetylcholine receptor", J Neurochem, 101 (2007) 1527-38.
• Hou X, Aguilar MI, Small DH. Transthyretin and familial amyloidotic polyneuropathy. FEBS J., 274 (2007) 1637-50.
• Losic D, Mechler A, Martin LL, Aguilar MI, Small DH, High resolution scanning tunnelling microscopy of the b-amyloid protein (Ab1-40) of Alzheimer's disease suggests a novel mechanism of oligomer assembly, J Struct Biol, 155 (2006) 104-10.
• Losic D, Martin LL, Aguilar MI, Small DH, "beta-Amyloid fibril formation is promoted by step edges of highly oriented pyrolytic graphite". Biopolymers, 84 (2006) 519-26.
• Aguilar MI, Small DH, ‘Surface Plasmon Resonance for the Analysis of b-Amyloid Interactions and Fibril Formation in Alzheimer's Disease Research' J Neurotox Res, 7 (2005) 17-27.
• Hou X, Richardson SJ, Aguilar MI, Small DH, "Binding of amyloidogenic transthyretin to the plasma membrane alters membrane fluidity and induces neurotoxicity", Biochemistry, 44 (2005) 11618-27.
• Fodero, L.R., Mok, S.S. Losic, D., Martin, L.L., Aguilar, M.I., Barrow, C.J., Livett, B.G. and Small, D.H. `a7-Nicotinic acetylocholine receptors mediate an Ab<sub>1-42</sub>-induced increase in the level of acetylcholineesterase in primary cortical neurons' J. Neurochem, 88 (2004) 1186-1193.
• Klug, G.M.G.A., Losic D., Subasinghe, S., Aguilar, M.I., Martin, L L. and Small D.H. ‘Ab oligomers induced my metal ions and low pH are distinct from those generated by slow spontaneous ageing at neutral pH', Eur J Biochem, 270 (2003), 4282-93.
• Subasinghe, S, Unabia, S., Barrow, C.J., Mok S.S., Aguilar M.I., Small D.H., `Cholesterol is necessary both for the toxic effect of Ab peptides on vascular smooth muscle cells and for Ab binding to vascular smooth muscle cell membranes', J. Neurochem, 203 (2003), 471-479.

Membrane Protein Purification & Membrane Proteomics

With the availability of the total human genome sequence, there is now an enormous effort directed towards the development of new technologies to allow the compositional and functional analysis of the corresponding proteome. It has been estimated that 30-40% of encoded DNA codes for membrane proteins and this class of proteins therefore represents a significant proportion of the cell’s complement of protein. However, membrane protein isolation and analysis continues to be an enormously challenging task despite the significant advances made in separation sciences over the last 30 years. The lack of robust separation techniques for membrane proteins has now led to a bottleneck in both their structural elucidation (there are only ~ 50 x-ray structures of membrane proteins compared to thousands of soluble protein structures) and high-throughput functional analysis. Furthermore, it has been estimated that approximately 70% of current drugs target membrane proteins, clearly demonstrating the importance of this class of proteins to the pharmaceutical industry.

We have developed new chromatographic materials, which are being applied to the isolation and analysis of membrane proteins with a particular focus on the proteomic analysis of a number of tissue sources.

Publications:

• Peng L, Kapp EA., Fenyö D, Kwon M-S, Jiang P, Wu S, Jiang Y, Aguilar MI, Ahmed N, Baker MS., Cai Z, Chen Y-J, Ch PV, Chung MCM, He F, Len ACL, Liao P-C, Nakamura K, Sai M N, Paik Y-K, Pan T-L, Poon TCW, Salekdeh H, Simpson RJ., Sirdeshmukh R. Srisomsap C, Svasti J, Tyan Y-C, Dreyer F, McLauchlan D, Rawson P and Jordan TW, 'Strategies for Membrane Proteomics: Preparation of the AOHUPO Membrane Proteomics Initiative Standard', Proteomics, 10 (2010) 4142-4148.
• Lee T-Z, Aguilar M.I., Protein separation using immobilized phospholipid chromatography', in Zachariou M (Ed) Methods in Molecular Biology: Affinity Chromatography: Methods & Protocols, Humana Press, 2008 Vol 421, p295-302.
• Davies M, Lee TH, Apffel A, Aguilar MI. "Hydrophobic and electrostatic forces control the retention of membrane peptides and proteins with an immobilised phosphatidic acid column", J Chromatogr, 1156 (2007) 167-173.
• Lee T-Z, Aguilar M.I., ‘Trends in the Development and Application of Functional Biomembrane Surfaces', Biotechnology Annual Reviews, 12 (2006) 85-136
• Lee T-H, Aguilar M.I., ‘HPLC of Peptides and Proteins', Encyclopedia of Molecular Cell Biology and Molecular Medicine, RA Meyers (Ed), Wiley NY, 2005, p245-295.
• Aguilar M.I. and Purcell, A.W., Analysis of peptides: Applications', Encyclopaedia of Analytical Science, 2005, Academic Press, Vol 7, p29-35
• Lee T-H and Aguilar M.I., `Biomembrane chromatography: Application to purification and biomembrane interactions'. Adv Chromatogr, 41 (2001) 175-201.
• Mozsolits H, Lee T-H, Wirth H-J, Perlmutter P and Aguilar M.I., `Interaction of peptide hormones with a phospholipid monolayer modified silica support', Biophys J, 77 (1999) 1428-1244.
• Lee T-H, Mozsolits H, and Aguilar M.I., `Measurement of the Affinity of Mellitin for Zwitterionic and Anionic Membranes Using Immobilised Lipid Biosensors', J. Pept. Res. 58 (2001) 464-476.
• Lee T-H, Rivett D, Werkmeister J, Hewish D and Aguilar M.I., `The interaction of amphipathic peptides with an immobilised model membrane', Lett in Pept Sci, 6 (1999) 371-380.

Nanotechnology in Food & Colloid Science

In the dairy industry, whey proteins have been widely used as functional ingredients in many food emulsions, including ice creams, salad dressings and frozen desserts. As effective emulsifiers, proteins not only adsorb to the interface, reducing the interfacial tension during the emulsification process, but also prevent droplet aggregation under various stresses that the product experiences during manufacture, transport, storage and usage. These interfacial activities of proteins are ultimately determined by their molecular structure and physicochemical properties. In collaboration with Dr Mary Ann Augustin and Dr Tim Wooster from CSIRO Food & Nutritional Sciences, we are applying a range of novel techniques to understanding the role of protein structure in food emulsions. The detection of specific substances in food is also of primary interst in quality control and safety. We have collaborated with Dr Yinqiu Wu and Dr Jing Yuan of Hort Research (NZ) in the development of novel high sensitivity biosensor-based assays for the detection of small molecules in various food items. The overall aim of these studies is to assist the development of new processes for emulsion-based food product manufacture.

Publications:

• Zhai J, Miles AJ, Pattenden LK, Lee TH, Augustin MA, Wallace BA, Aguilar MI and Wooster TJ, ‘The use of synchrotron radiation circular dichroism to characterize b-lactoglobulin structures at the oil-water interface of emulsions', Biomacromolecules, 11(2010) 2136-2142.
• Yuan J, Deng, D, Lauren, D, Aguilar MI, Wu Y, "Surface plasmon resonance biosensor for the detection of ochratoxin A in cereals and beverages". Anal Chim Acta, 656 (2009) 63-71.
• Yuan J, Addo, J, Aguilar M I, Y Wu, ‘Continuous high affinity surface plasmon resonance assay for chloramphenicol'. Anal Biochem, 390 (2009) 97-99.
• Yuan J, Oliver R, Aguilar M I, Y Wu, ‘Surface plasmon resonance assay for chloramphenicol'. Anal Chem, 80 (2008) 8329-8333.
• Yuan J, Oliver R, Li J, Lee TH, Aguilar M I, Y Wu, ‘Nanogold particles for sensitivity enhancement of SPR assay of progesterone based on mixed self-assembled monolayers'. Biosens and Bioelect, 23 (2007) 144-148.

PEPTIDOMIMETIC DRUG DESIGN

The use of peptidomimetics has emerged as a powerful means for overcoming the limitations inherent in the physical characteristics of peptides thus improving their therapeutic potential. A peptidomimetic approach that has emerged in recent years with significant potential, is the use of ß-amino acids. ß-Amino acids are similar to ß-amino acids in that they contain an amino terminus and a carboxyl terminus. However, in ß-amino acids two carbon atoms separate these functional termini. ß-amino acids, which results in a total of 4 possible diastereoisomers for any given side chain. The flexibility to generate a vast range of stereo- and regioisomers, together with the possibility of disubstitution, significantly expands the structural diversity of ß-amino acids thereby providing enormous scope for molecular design. The incorporation of ß-amino acids has been successful in creating peptidomimetics that not only have potent biological activity, but are also resistant to proteolysis and we are applying these techniques to a range of protein targets.

Therapeutic Peptidase Inhibitors and Receptor Ligands

The action of most neuropeptides is terminated by specific extracellular peptidases and these enzymes therefore play an important role in the regulation of the function of the central nervous system.  The availability of inhibitors of these enzymes is important for characterising the role of these enzymes in peptide signaling in the brain and ultimately for the development of new therapeutic agents for the treatment of cardiovascular disease.  In collaboration with Prof Patrick Perlmutter (School of Chemistry), Prof Ian Smith, together with Prof Rob Widdop (Dept of Pharmacol) and Dr Emma Jones (Dept of Pharmacol) and Prof David Small (Menzies Research Institute) we are focusing on a number of enzyme and receptor targets. 

EC 3.4.24.15 (EP 24.15) is a widely-distributed enzyme involved in the regulation of blood pressure. The specific function of this enzyme is unknown, but it has been implicated in the metabolism of bradykinin.

Membrane-bound aminopeptidase P (AP-P) also participates in the degradation of bradykinin in several vascular beds. Together with angiotensin-converting enzyme, AP-P is responsible for a large proportion of the breakdown of bradykinin. Since bradykinin exhibits potent vasodilatory and cardioprotective effects, there is a therapeutic benefit to inhibiting these enzymes and increasing endogenous levels of bradykinin.

ACE2 is a very recently discovered enzyme and is expressed largely in the kidney and heart suggesting important functions in cardiovascular and renal systems and currently there is an enormous interest in this enzyme as it has been proposed to be an essential regulator of heart function in vivo.

BACE-1 is an enzyme involved in the production of b-amyloid, the protein implicated in Alzheimer's disease (AD). Inhibitors of this enzyme may be useful in the treatment of AD.

We are currently designing novel peptide and peptidomimetic based inhibitors of these enzymes together with related GPCRs in order to  develop more effective approaches for the treatment of cardiovascular disorders and Alzheimer's diesease.

The proposed mechanism of action of ECE, 24.15 & 24.16. These sites are targets for new drugs.

Publications:

• Clayton DJ, Hanchapola I, Hausler N, Unabia S, Widdop RE, Smith AI, Perlmutter P and Aguilar MI, ‘β-Amino acid substitution to investigate the recognition of angiotensin II (AngII) by angiotensin converting enzyme 2 (ACE2)', J Mol Recog. 24 (2011) 235-244.
• Bergman YE, Del Borgo MP, Devi Gopalan R, Jalal S, Unabia SE, Ciampini M, Clayton DJ, Fletcher JM, Mulder RJ, Wilce JA, Aguilar MI and Perlmutter P, ‘The synthesis of stapled b³-peptides by ring closing metathesis'. Org Lett., 11 (2009) 4438-4440.
• Bergman Y, Ciampini M, Jalal S, Lagiakos HR, Aguilar MI and Perlmutter P, The synthesis of Fmoc-O-Allyl b-serine. Tetraherdron Assymetry, 19 (2009) 2861-2863.
• Lagiakos HR, Aguilar MI, Perlmutter P, "A mild method for the efficient, [3,3]-sigmatropic rearrangement of N,O-diacyl hydroxylamines", J Org Chem, 74 (2009) 8001-8003.
• Aguilar MI, Purcell AW, Devi, R, Lew, R, Purcell AW, Rossjohn J, Smith AI, and Perlmutter P. "b-Amino acid-containing hybrid peptides - new opportunities in peptidomimetics". Organic and Biomolecular Chemistry, 5 (2007) 2884-90.
• Aguilar MI, Fallon GD, Mayes P, Nordin S, Robinson AJ, Rose ML, Vounatsos F, Wilman B, Perlmutter P, The Asymmetric Imino-Aldol Approach To The Enantioselective Synthesis Of Beta-Amino Acids. Lett. Pep. Sci., 10 (2004), 597-604.
• Steer DL, Lew R, Perlmutter P, Smith AI and Aguilar MI, `b-Amino acids: Versatile peptidomimetics', Curr. Med. Chem. 9 (2002) 811-822.
• Steer DL, Lew R A, Perlmutter, P, Smith AI and Aguilar MI, `Inhibitors of metalloendopeptidase E.C. 3.4.24.15 and EC 3.4.24.16 stabilised against proteolysis by the incorporation of b-amino acids', Biochemistry, 41 (2002) 10819-10826.
• Steer DL, Lew R, Perlmutter P, Smith AI and Aguilar MI, `The Use of b-Amino Acids in the Design of Peptidase and Protease Inhibitors' Lett in Pept Sci, 8 (2002) 241-246.
• Lew R A, Boulos, E, Stewart K M, Perlmutter P, Harte M, Bond S, Gerryn, S, Norman,M U, Lew M J, Aguilar M I, Smith A I, `Substrate Analogs Incorporating b-Amino Acids: Potential Use in Peptidase Inhibition', FASEB J. 15 (2001) 351-356.

Peptide-Based Vaccines - High Affinity Peptide Ligands for Class I MHC Proteins

Class I major histocompatibility complex (MHC) proteins play a key role in immune surveillance by selectively binding to intracellular peptide antigens and presenting them at the cell surface to cytotoxic T-lymphocytes (CTL). Interference of this process by analogues of peptide antigens has been shown to cause significant changes in T cell function which suggests that these analogues have significant potential as immunotherapeutic agents. However, the challenge has been to rationally design analogues of peptide antigens which cause subtle changes in antigen recognition. In collaboration with Assoc Prof Tony Purcell at the Dept of Biochemistry & Molecular Biology at the University of Melbourne, we are currently using our peptidomimetic approaches to the design of novel T-cell antagonists.

Graphic illustration of peptide antigens presented by the MHC class I heavy chain. Alteration of such antigenic peptide structure can affect the T-cell recognition and therefore the activation of CD8+ CTL.

Publications:

• Purcell AW, Dudek NL, Perlmutter P, Aguilar MI, Williamson N, ‘Peptide based vaccines: from discovery to application', Curr Pharm Design, 16 (2010) 3149-3157.
• Webb AI, Dunstone MA, Williamson NA, Price JD, de Kauwe A, Chen W, Oakley A, Perlmutter P, McCluskey J, Rossjohn J, Aguilar MI, Purcell AW ‘T cell determinants incorporating β-amino acid residues are protease resistant and remain immunogenic in vivo'. J Immunol, 175 (2005) 3810-18.
• Webb AI, Dunstone MA, Chen W, Aguilar MI, Chen Q, Jackson H, Chang L, Kjer-Nielsen L, Beddoe T, McCluskey J, Rossjohn J, Purcell AW, ‘The structure of HLA A2 complexed to peptides related to the tumor antigen NY-ESO-1 and the rational design of a new immunogenic analogue', J Biol Chem (2004) 279:23438-46.
• Webb AI, Borg N, Dunstone MA, Kjer-Nielsen L, Beddoe T, McCluskey J, Carbone F, Bottomley SP, Aguilar MI, Purcell AW, Rossjohn J, ‘T cell repertoire selection and anti-viral resistance associated with a conformational switch in polymorphic H-2K molecules', J Immunol, (2004) 173:402-9.
• Webb AI, Aguilar MI, Purcell AW, ‘Optimisation of peptide-based cytotoxic T cell determinants using non-natural amino acids'. Lett. Pep. Sci., 10 (2004), 561-569

Novel Targets for the Treatment of Cardiovascular Disease

This project aims to understand the cardiovascular system and how certain systems such as the renin angiotensin system exhibit markedly increased activity which contributes to a number of cardiovascular diseases and premature ageing.
In particular, overactivity of the main effector hormone, angiotensin II, contributes to hypertension, coronary heart disease and atherosclerosis, as well as heart failure and stroke. These are major chronic diseases that account for approximately 40% of all-cause mortality in the western world. Fortunately, these diseases are generally managed with medicines for the rest of the patient's life. However, many patients are resistant to current treatments and often require multiple medications, therefore, validation of novel therapeutic targets is vital to combat life-threatening chronic diseases of the cardiovascular system.
We are examining novel components of the renin angiotensin system that act as ‘physiological brakes' against the excitatory effects of angiotensin II. We are particularly looking at various angiotensin receptor isoforms as potential therapeutic targets that can treat these chronic cardiovascular diseases.

Publications:

• Jones ES, Del Borgo MP, Kirsch JF, Clayton DJ, Bosnyak S, Welungoda I, Hausler N, Unabia S, Perlmutter P, Thomas WG, Aguilar MI, Widdop RE, ‘A single beta-amino acid substitution to angiotensin II confers remarkable AT2 receptor selectivity and vascular function', Hypertension, 57 (2011) 570-576.
• Bosnyak S, Jones ES, Christopoulos A, Aguilar MI, Thomas WG and Widdop RE, ‘Relative affinity of angiotensin peptides and novel ligands at AT1 and AT2 receptors', Clinical Sciences (2011) in press.

GENERAL PROJECT AREAS

1. G-protein-coupled receptor regulation (with Prof Wally Thomas [University of Queensland])
2. Role of the membrane in protein misfolding - application to Alzheimer's disease (with Prof David Small, Menzies Research Institute, Tasmania).
3. The design and synthesis of novel peptide inhibitors involved in cardiovascular disease (with Prof Patrick Perlmutter [Dept of Chemistry], Dr Mark DelBorgo, Prof Rob Widdop (Pharmacology) and Dr Emma Jones (Pharmacology).
4. The mechanism of action of antimicrobial peptides (with Prof Frances Separovic, Uni of Melbourne) and Dr John Lee.
5. The design and synthesis of novel anti-HIV compounds (with Assoc Prof Gilda Tachjdian, [Burnet Institute] and Dr Miranda Shehu-Xhilaga).
6. Design of peptide-based vaccines (with Assoc Prof Tony Purcell [Univ. of Melbourne])