Dr Terry Mulhern
Position: Russell Grimwade Fellow
Affiliation: Department of Biochemistry and Molecular Biology, University of Melbourne
The Russell Grimwade School of Biochemistry and Molecular Biology
The University of Melbourne
Melbourne, Vic 3010
Phone: +61 (03) 8344 5931
Fax: +61 (03) 9347 7730
Signal Transduction and Human Diseases:
Understanding Structure and Function
Signal transduction is a complex web of molecular recognition events and activation and deactivation utilising molecular switches such as phosphorylation and conformational change. An understanding of signal transduction is itself important, but is made more so by the potential for disease states to develop when it goes wrong e.g. cancer and diabetes. The projects in my group are focused on developing an understanding of how signal transduction defects lead to human diseases. The techniques employed in these projects include: recombinant DNA manipulation, protein expression and purification, activity/binding assays and biophysical analyses including circular dichroism (CD), analytical ultra centrifugation, calorimetry, protein crystallography and in particular, nuclear magnetic resonance (NMR) spectroscopy.
Protein structure and interactions by NMR spectroscopy
NMR spectroscopy provides a window into molecular structure, interactions and dynamics at an atomic level. There have been some spectacular developments in NMR techniques in recent years, which have allowed us to determine the 3-dimensional solution structure of biologically important molecules of ever increasing size and complexity. NMR also provides techniques for studying protein dynamics and the interaction between biopolymers, be they protein or nucleic acids, and their ligand partners in solution. These features have made NMR spectroscopy a key discipline in many areas of biomedical research and the pharmaceutical industry. Researchers within the Department have access to Varian 400, 500 and 600 MHz spectrometers and a suite of workstations for data analysis and other computational needs. Dr Paul Gooley leads another NMR group giving the Department critical mass in areas such as pulse sequence development, isotopic labelling, triple-resonance spectroscopy and protein structure calculation.
Current Research Projects:
Protein-protein interactions of the ATM kinase
Ataxia telangiectasia (A-T) is an autosomal recessive disease with a complex phenotype characterized by progressive neurological abnormalities, premature aging and a predisposition to cancer. It is estimated that it affects 1 in 40,000 births. The disease arises from mutations to the ATM (mutated in A-T) gene, which encodes a PI3K-like Ser/Thr kinase. When ionising radiation damages DNA ATM localises to double strand breaks where it phosphorylates proteins stimulating DNA repair mechanisms. We are particularly interested in defining the interacting regions of ATM and its substrates p53, which is mutated in the majority of human tumours, and BRCA1, which is mutated in many breast and ovarian cancer patients. In collaboration with Dr Kum Kum Khanna (Queensland Institute of Medical Research) and Dr Melissa Brown (University of Queensland).
Regulation of the Src family enzymes
The Src family of protein tyrosine kinases (SFKs) are important intracellular signalling enzymes. They are involved in many processes including cell proliferation and differentiation and as such are proto-oncogenes. SFKs share a similar domain structure being composed of a Src homology 3 (SH3) and a Src homology 2 (SH2) domain followed by the catalytic subunit. Each family member also contains a unique N-terminal region ranging in length from 60 to 80 residues. These unique regions share very little sequence similarity and are thought to play a major role in mediating the different activities and localisations of the SFKs. While the structure and interactions between the Src homology and catalytic domains are relatively well understood, the same cannot be said for the unique regions. We are interested in analysing the role of domain-domain and domain-ligand interactions in the regulation of SFK activity. In collaboration with Dr H. -C. Cheng (University of Melbourne), Professor Iain Campbell and Dr Martin Noble (University of Oxford).
Activating mutants of the IL-3/IL-5 and GM-CSF receptors
Granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3) and interleukin-5 (IL-5) are haemopoietic cytokines that bind to cell-surface receptors that share a common polypeptide-chain (the beta-chain). The beta-chain is the critical signalling subunit of the receptor and its fourth domain plays a critical role in interactions with ligands and also contains residues whose mutation can lead to ligand-independent activation of the receptor. We are interested in elucidated of the structural features of these mutants that lead to factor-independence. In collaboration with Dr Christopher Bagley and Dr Richard D'Andrea (Hanson Centre For Cancer Research, Adelaide).
- Mulhern TD, Howlett GJ, Reid GE, Simpson RJ, McColl DJ, Anders RF & Norton RS. (1995) Solution structure of a polypeptide containing four heptad repeat units from a merozoite surface antigen of Plasmodium falciparum. Biochemistry 34, 3479-91. (Impact Factor: 4.06; Citations: 16)
- Mulhern TD, Shaw GL, Morton CJ, Day AJ & Campbell ID. (1997) The SH2 domain from the tyrosine kinase Fyn in complex with a phosphotyrosyl peptide reveals insights into domain stability and binding specificity. Structure 5, 1313-23. (Impact Factor: 6.03; Citations: 14)
- Mulhern TD, Booker GW & Cosgrove L. (1998) A third fibronectin-type-III domain in the insulin-family receptors. Trends Biochem. Sci. 1998 23, 465-6. (Impact Factor: 14.40; Citations: 14)
- Chung EW, Henriques DA, Renzoni D, Morton CJ, Mulhern TD, Pitkeathly MC, Ladbury JE & Robinson CV. (1999) Probing the nature of interactions in SH2 binding interfaces -evidence from electrospray ionization mass spectrometry. Protein Sci. 8, 1962-70. (Impact Factor: 3.55; Citations: 11)
- Mulhern TD, Lopez AF, D'Andrea RJ, Gaunt C, Vandeleur L, Vadas MA, Booker GW & Bagley CJ. (2000) The solution structure of the cytokine-binding domain of the common beta-chain of the receptors for granulocyte-macrophage colony-stimulating factor, interleukin-3 and interleukin-5. J. Mol. Biol 297, 989-1001. (Impact Factor: 5.36; Citations: 7)
- Chia BC, Carver JA, Mulhern TD & Bowie JH. (2000) Maculatin 1.1, an antimicrobial peptide from the Australian tree frog, Litoria genimaculata: solution structure and biological activity. Eur. J. Biochem. 267, 1894-1908. (Impact Factor: 3.00; Citations: 12)
- Arold ST, Ulmer TS, Mulhern TD, Werner JM, Ladbury JE, Campbell ID & Noble MEM. (2001) The role of the SH3-SH2 interface in the regulation of Src kinase. J Biol Chem. 276, 17199-17205. (Impact Factor: 6.70; Citations: 8)
- Hatters DM, Wilson L, AtclImpact Factorfe B, Mulhern TD, Guzzo-Pernell N & Howlett GJ. (2001) Sedimentation analysis of novel DNA structures formed by homo-oligonucleotides. Biophys J. 80, 371-381. (Impact Factor: 4.64; Citations: 3)
- Khanna KK, Lavin MF, Jackson SP & Mulhern TD. (2001) ATM, a central controller of cellular responses to DNA damage. Cell Death Differ. 8, 1052-1065. (Impact Factor: 5.70; Citations: 23)
- Pursglove SE*, Mulhern TD*, Mackay JP, Hinds MG & Booker GW. (2002) The solution structure and intramolecular associations of the Tec kinase Src homology 3 domain. J Biol Chem. 277, 755-762. (*equal contribution) (Impact Factor: 6.70; Citations: 6)
- High field NMR: 800, 600 & 500 MHz facilities
- Steady state and time resolved fluorescence spectroscopy
- Circular dichroism spectroscopy
- Analytical ultracentrifugation
- peptide synthesis
- amino acid sequencing
Dr Jill Trewhella, Los Alamos National Laboratory, New Mexico USA