Peptide Laboratory
Laboratory Head
Prof John D Wade BSc (Hons) PhD FRACI FRSC
Contact Details
Email: | |
Phone: | +61 (0)3 8344 7285 |
Fax: | +61 (0)3 9348 1707 |
Number of
Staff: 5
- Dr Eleni Giannakis eleni.giannakis@florey.edu.au
- Dr Akhter Hossain akhter.hossain@florey.edu.au
- Dr Suode Zhang suode.zhang@florey.edu.au
- Mr Feng Lin feng.lin@florey.edu.au
- Mrs Mary Macris mary.macris@florey.edu.au
Students: 3
- Alessia Belgi
- Linda Chan
- Fazel Shabanpoor
Research Interests
The principal focus of our laboratory is the structure and function relationship of members of the insulin superfamily, particularly relaxins-2 and 3 and insulin-like peptide 3 (INSL3). We are endeavouring to understand how their three dimensional shape dictates their unique biological properties and to use this information to design and construct mimetics of these peptides that may have therapeutic applications.
Relaxin and INSL3 each consists of two peptide chains, A and B, that are held together by three disulfide bonds, one within the A-chain and two intermolecular between the two chains. In humans, there are two relaxins known as 2 and 3. Relaxin-2 is produced principally in the ovaries of preganant females and has important post-partum roles. Relaxin-3 was recently discovered at the Florey and appears to be a neuropeptide having key roles in stress and appetite regulation. INSL3 has a primary role in initiating testes descent in the developing male and in the adult is an important component of the gonadotropic axis and regulates germ cell maturation.
We use state-of-the-art peptide synthesis technology to prepare these peptide hormones in sufficient quantities to allow their detailed biochemical study, that is, how they bind to their receptors to cause their unique biological effects. We also use drug design techniques in conjunction with tertiary structural techniques to prepare analogues of the peptides that are predicted to be more potent, longer lasting and specific.
We also undertake detailed protein profiling and biomarker discovery with the goal of using this technology to study the role of these peptides in health and disease. Towards this aim, use a Ciphergen surface-enhanced laser desorption/ionization (SELDI) ProteinChip System©. It is a multi-dimensional protein recognition tool and structure analyser. It enables protein capture, purification, analysis, and processing from complex biological mixtures directly on derivatised surfaces. Peptides and proteins are affinity-immobilised onto the surface of chemically or biochemically activated ProteinChips. Retained analytes are detected according to molecular weight and quantitated by the use of a time-of-flight mass spectrometer (TOF MS) equipped with a laser desorption ion source. We have also been using this technology to understand the role of amyloid-beta in neurological disease and have logically extended the work to include other areas of neuroscience.
Current Projects
- Determining the structure and function relationships of human relaxin-2 and -3 and insulin-like peptide 3 (Akhter Hossain, Feng Lin, Fazel Shabanpoor, Suode Zhang, John Wade with Tony Hughes, Department of Pharmacology, University of Melbourne).
- Protein profiling of relaxin KO mice, a model of progressive fibrosis (Eleni Giannakis, Chrishan Samuel, Mary Macris, Geoffrey Tregear, John Wade).
- Capture and quantitation of Abeta, a marker for Alzheimer's Disease (Eleni Giannakis, John Wade and Lin Hung with Kevin Barnham, Keyla Perez and Colin Masters, Department of Pathology, University of Melbourne).
- The mechanism of binding of Abeta to cell membranes (Keyla Perez with Frances Separovic, Bio21 Institute, University of Melbourne).
- Capture and quantitation of alpha-synuclein, a marker for Parkinson’s Disease (Eleni Giannakis and John Wade with Rogan Tinsley, Janelle Nunan and Mal Horne).
- Protein profiling of D2 KO mice, a model of Parkinson’s Disease (Eleni Giannakis, Mary Macris and John Wade with Janelle Nunan, Rogan Tinsley and Mal Horne).
- Design and development of novel antibacterial peptides (John Wade and Feng Lin with Laszlo Otvos, Temple University, Philadelphia, USA, and Ralf Hoffman, Leipzig University, Germany).
- Mining reptile venom for new peptide therapeutics (John Wade and Feng Lin with Bryan Greig Fry, Australian Venom Research Unit, University of Melbourne).
- The structure and function of insulin analogues as potential new diabetes treatments (Akhter Hossain, Feng Lin, John Wade with Andrea Robinson, School of Chemistry, Monash University).
Laboratory Techniques
- Solid phase peptide synthesis
- Reversed phase-HPLC
- MALDI-TOF and SELDI-TOF mass spectrometry
- Computer-aided drug design
- Amino acid analysis
- Peptide engineering
Funding
- National Health and Medical Research Council (Australia) - Project grants
- Australian Research Council Discovery grant
- Diabetes Australia
- The Ian Potter Foundation
- The R.E. Ross Foundation
Additional Information
Recent Peptide Laboratory publications
Rosengren, K.J., Zhang, S., Lin, F., Daly, N.L., Scott, D.J., Hughes, R.A., Bathgate, R.A.D., Craik, D.J. and Wade, J.D. (2006) Solution structure and characterization of the receptor binding surface of insulin-like peptide 3. Journal of Biological Chemistry, 281, 28287-28295.
Samuel, C.J., Lin, F., Zhao, C., Layfield, S.L., Hossain, M.A., Zhang, S., Giannakis, E., Bathgate, R.A., Tregear, G.W. and Wade, J.D. (2007) Improved chemical synthesis and demonstration of relaxin receptor binding activity and biological activity of mouse relaxin. Biochemistry, 46, 5374-5381.
Kuei, C., Sutton, S., Bonaventure, P., Pudiak, C., Shelton, J., Zhu, J., Nepomuceno, D., Wu, J., Chen, J., Kamme, F., Seierstad, M., Hack, M.D., Bathgate, R.A.D., Hossain, M.A., Wade, J.D., Atack, J., Lovenberg, T.W. and Liu, C. (2007) R3(BΔ23-27)R/I5 chimeric peptide, a selective antagonist for GPCR135 and GPCR142 over LGR7: In vitro and in vivo characterization. Journal of Biological Chemistry, 282, 25425-25435, 2007.
Lau, T-L., Gehman, J. D., Wade, J.D., Masters, C.L., Barnham, K.J. and Separovic, F. (2008) Membrane interactions and the effect of metal ions of the amyloidogenic fragment Aß(25-35) in comparison to Aß(1-42). Biochimica et Biophysica Acta-Biomembranes, 1768, 2400-2408.
Svendsen, A.M., Vrecl, M., Ellis, T.M., Heding, A., Kristensen, J.B, Wade, J.D., Bathgate, R.A.D., De Meyts, P. and Nøhr, J. (2008) Cooperative binding of insulin-like peptide 3 to a dimeric relaxin family peptide receptor 2. Endocrinology, 149, 1113-1120.
Giannakis, E., Pacifico, J., Smith, D.P., Hung, L-W., Master, C.L., Tregear, G.W., Cappai, R., Wade, J.D. and Barnham, K.J. (2008) Dimeric structures of α-synuclein bind preferentially to lipid membranes. Biochimica et Biophysica Acta - Biomembranes 1778, 1112-1119.
Hossain, M.A., Bathgate, R.A., Kong, C., Shabanpoor, F., Zhang, S., Haugaard-Jönsson, L.M., Rosengren, K.J., Tregear, G.W. and Wade, J.D. (2008) Synthesis, conformation and receptor binding activity of human insulin-like peptide 5 (INSL5). ChemBioChem, 9, 1816-1822.
Hossain, M.A., Rosengren, K.J., Haugaard-Jönsson, L.M., Zhang, S., Layfield, S., Ferraro, T., Daly, N.L., Tregear, G.W., Wade, J.D. and Bathgate, R.A. (2008) The A-chain of human relaxin family peptides has distinct roles in the binding and activation of the different relaxin family peptide receptors. Journal of Biological Chemistry, 283, 17287-17297.
Shabanpoor, F., Hughes, R.A., Bathgate, R.A.D., Zhang, S., Scanlon, D.B., Lin, F., Hossain, M.A., Separovic, F. and Wade, J.D. Solid-phase synthesis of europium-labelled human INSL3 as a novel probe for the study of ligand-receptor interactions. (2008) Bioconjugate Chemistry, 19, 1456-1463.
Otvos, L. Jr., Terrasi, M., Cascio, S., Cassone, M., Knappe, D., Stawikowski, M., Cudic, P., Wade, J.D., Hoffmann, R. and Surmacz, E. (2009) Development of a pharmacologically improved peptide agonist of the leptin receptor. Biochimica et Biophysica Acta – Molecular Cell Research, 1783, 1745-1754.
Svendsen, A.M., Zalesko, A., Kønig, J., Vrecl, M., Heding, A., Kristensen, J.B., Wade, J.D., Bathgate, R.A., De Meyts, P. and Nøhr, J. (2008) Negative cooperativity in H2 relaxin binding to a dimeric relaxin family peptide receptor 1. Molecular and Cellular Endocrinology, 296, 10-17.
Haugaard-Jönsson, L.M., Hossain, M.A., Daly, N.L., Bathgate, R.A., Wade, J.D., Craik, D.J. and Rosengren, K.J. (2008) Structure of the R3/I5 chimeric relaxin peptide, a selective GPCR135 and GPCR142 agonist. Journal of Biological Chemistry, 283, 23811-23818.
Hossain, M.A., Bathgate, R.A.D., Rosengren, K.J., Shabanpoor, F., Tregear, G.W. and Wade, J.D. (2009) The structural and functional role of the B-chain C-terminal arginine in the relaxin-3 peptide antagonist, R3(BΔ23-27)R/I5. Chemical Biology & Drug Design, 73, 46-52.
Shabanpoor, F., Hughes, R.A., Zhang, S., Bathgate, R.A.D., Layfield, S., Hossain, M.A., Tregear, G. W., Separovic, F. and Wade, J.D. (2009) Effect of helix-promoting strategies on the LGR8 receptor activity of novel analogues of the B-chain of human insulin-like peptide 3, INSL3. Amino Acids (in press).
Hossain, M.A., Rosengren, K.J., Bathgate, R.A., Tregear, G.W., van Lierop, B.J., Robinson, A.J. and Wade, J.D. (2009) Solid phase synthesis and structural activity of novel dicarba analogues of human relaxin-3 (INSL7) that exhibit full biological activity. Organic and Biomolecular Chemistry (in press).
Haugaard-Jönsson, L.M., Hossain, M.A., Daly, N.L., Craik, D.J., Wade, J.D. and Rosengren, K.J. (2009) Structure of human insulin-like peptide 5 and characterization of conserved hydrogen bonds and electrostatic interactions within the relaxin framework. Biochemical Journal (in press).
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