Summer Internships 2012

Internship projects available for Summer 2012

Understanding protein misfolding of TDP-43 in amyotrophic lateral sclerosis(ALS) and frontotemporal dementia (FTD)

Protein misfolding can result in protein aggregation, a phenomenon associated with a wide range of devastating neurodegenerative conditions such as Alzheimer’s, Parkinson’s, Fronto-temporal dementia (FTD) and amyotrophic lateral sclerosis (ALS), The TAR DNA binding protein (TDP-43) is the major component of cytoplasmic inclusions found in 90% of ALS and 70% of FTD patients. TDP-43, A highly conserved 43kDa multi-domain nuclear protein, predominantly located in the nucleus that regulates RNA transcription, splicing and transport. In disease, it mislocalizes to the cytoplasm where it aggregates to form hallmark pathological inclusions.

The aim of this project is to obtain a molecular understanding of the aggregation pathway of TDP-43 and thereby develop therapeutic strategies to inhibit the process. This will be in collaboration with the leading ALS group headed by Prof Christopher E Shaw,. His group are pioneering the ALS field (see ref 1-3). They have shown that mutation of the gene encoding TDP-43 results in increased protein fragmentation in transfected neurons in culture and is neurotoxic in the chick embryo.

In this project we will study at a residue-specific level the structural and folding properties of TDP-43, and disease associated variants, using structural biology, in particular biophysics and NMR spectroscopy together with biochemical techniques.

John Christodoulou (JC) is a lecturer at the joint Birkbeck/UCL Institute of Structural and Molecular Biology and Director of the Centre for NMR Spectroscopy. His research group’s activities focus on protein folding, misfolding and aggregation including during biosynthesis on the ribosome as well as to characterize the Parkinson’s disease associated protein, α-synuclein which aggregates to form amyloid deposits in Parkinson’s disease.

Christopher E Shaw is Professor of Neurology and Neurogenetics, and Director of the MRC Centre for Neurodegeneration Research at King’s College, London. His group study the genetics and pathobiology of ALS and FTD.

References:

1. TDP-43 mutations in familial and sporadic amyotrophic lateral sclerosis. Sreedharan J, et al, Science (2008) 319, 1668-72.
2. Vance C, et al. Mutations in FUS, an RNA processing protein, cause familial amyotrophic lateral sclerosis type 6. Science. 2009;323:1208-11.
3. Nuclear import impairment causes cytoplasmic trans-activation response DNA-binding protein accumulation and is associated with frontotemporal lobar degeneration Nishimura et al, Brain (2010) 133, 1763-71.
4. Mapping long-range interactions in a-synuclein using spin-label NMR and ensemble molecular dynamics simulations Dedmon et al. J. Am. Chem. Soc. (2005) 127, 476-477.
5. Chaperone Proteostasis in Parkinson’s disease: Stabilization of the Hsp70/ -synuclein Complex by Hip Roodveldt, C Bertoncini, C.W., Andersson, A., van derGoot, et al. Embo. J. (2009) 28, 3758-70
6. On the mechanism of non-specific inhibitors of protein aggregation: dissecting the interactions of a-synuclein with Congo red and LacmoidLendel, C., Bertoncini, C.W.et al. Biochemistry (2009) 48, 8322-34.
7. Probing ribosome-nascent chain complexes produced in vivo by NMR spectroscopy Cabrita, L.D., et al. P.N.A.S (2009) 106, 22239-44.

Project supervisor/ host lab
Dr John Christodoulou, SMB, University College London

Analysis of mutations causing a dominantly inherited, complex immune disorder

In our collaborative project based on genetic analysis of families with a unique combination of autoimmunity and immunodeficiency we identified mutations within PLCG2 gene encoding one of the key second messenger producing enzymes, Phospholipase C. These mutations co-segregated with disease and affected the enzyme function. Our first aim is to extensively characterize enzymes harboring mutations. We will analyze their catalytic properties in vitro and the effects following introduction of these variants into different cell types from the immune system. Secondly, we plan structural, NMR studies of the isolated domains affected by the mutations to understand changes underpinning differences in the enzyme function.

The student will participate in experimental approaching addressing the first aim and will gain experience in cloning, cell transfection and some aspects of microscopy.

Project supervisor/ host lab
Dr Matilda Katan, Molecular Mechanisms of Signalling Team, SMB, UCL

Students from all Biological disciplines will be considered for this project.

Cloning and expression of kinesin motors from the malaria parasite

-- This project has now been filled and is no longer available --

Malaria is caused by the intracellular Plasmodium parasite and infects over 300 million people a year, killing more than 1 million. The complex life cycle of the parasite, the small number of drugs available and emerging drug resistance mean that novel drug targets are desperately needed. Kinesins are ATP-dependent microtubule-based molecular motors and are potential targets for anti-malarial drugs. The overall aim of this project is to study the motor domains of kinesin from malaria. The motor domain from a malaria kinesin will be cloned for recombinant expression in E. coli. If successful, protein purification and biochemical characterisation will be performed. In particular, kinesin ATPase activity and the way the motors interact with microtubules in vitro will be studied and can provide considerable insight into their functions in vivo. These studies will contribute insight into kinesin function in malaria and help identify suitable targets for screening of small molecule inhibitors.

Outcomes
Methods for cloning genes for recombinant expression in bacteria; optimisation of protein expression in bacteria; strategies for purifying recombinant protein from bacteria (if possible); assays to measure kinesin activity (if possible).

References
Morrissette NS, Sibley LD (2002) Cytoskeleton of apicomplexan parasites. Microbiol Mol Biol Rev 66, 21-38.

Miki H, Okada Y, Hirokawa N (2009) Analysis of the kinesin superfamily: insights into structure and function. Trends Cell Biol 15, 467-76 Deadline for enquiries: Jan 6th, 2012

Supervisor/ host lab
Dr Carolyn Moores, Department of Biological Sciences, Birkbeck

Deadline for enquiries
6 January 2012

Interactions of complement factor H with its ligands

-- This project has now been filled and is no longer available --

Age-related macular degeneration (AMD) is the main cause of blindness in the Western World in people over 55 years. Deposition of proteins, ions and lipids as "drusen" between the blood circulation and the retina is a major risk factor in AMD. Drusen usually grow in size and spread with age, disrupting the normal function of the retina, but may disappear. We recently discovered that the interaction between C-reactive protein and complement factor H may be a crucial factor in AMD (“Paper of the Week” in J.Biol.Chem. in January 2010). In order to prove further the involvement of intact complement factor H with this (or other ligands), the summer project will involve the purification of the homozygous high-risk and low-risk forms of factor H from patient plasma, then measurements will be made using surface plasmon resonance to define the conditions under which Factor H will interact with C-reactive protein (or other ligands) on sensor surfaces to which Factor H or C-reactive protein has been immobilised.

Supervisor/ host lab
Professor Steve Perkins, Structural Immunology Group. Molecular Interactions Facility

Students for this project will ideally be currently undertaking study in Biochemistry and/or Immunology

Deadline for enquiries
15 January 2012

CDC37 Dephosphorylation - A key step in the regulation of HSP90 Function

-- This project has now been filled and is no longer available --

Hsp90 is an essential molecular chaperone that contributes to the activation of over 50 kinases in the cell. Their association with Hsp90 is dependent on the cochaperone Cdc37, which recruits these kinases to Hsp90. The phosphorylation status of Cdc37 is one factor known to regulate this activity. This project uses crystallography, biochemical and biophysical approaches to investigate the molecular mechanism of dephosphorylation of Cdc37 since this step is essential for the activation of kinases dependent on Hsp90. Students will gain experience of protein expression and purification, crystallization and biochemical assays to investigate dephosphorylation.

Supervisor/ host lab
Dr Cara Vaughan, ISMB Lecturer in Structural Biology, Department of Biological Sciences, Birkbeck

This project is suitable for all Biosciences undergraduates

Deadline for enquiries
16 January 2012

Mapping the interactions of Hsp90 in the assembly of the yeast kinetochore

-- This project has now been filled and is no longer available --

The molecular chaperone Hsp90 is an essential protein involved in many signaling pathways and is a drug target for several diseases including cancer. The cochaperone Sgt1 recruits several proteins involved in kinetochore assembly and function to Hsp90 for activation. The kinetochore is the complex of proteins that bridges sister chromatids and microtubules in mitosis; it is essential for fidelity of genetic propagation during cell division. This project aims to contribute to a molecular description of the mechanisms involved in kinetochore assembly by focusing on the protein Ctf13, a protein of the yeast kinetochore that requires Hsp90 and Sgt1 function for its activation and the subsequent assembly of a functional kinetochore. The goal of this project is to understand how Hsp90 and Sgt1 facilitate the activation of Ctf13. This project will use crystallography and biochemical approaches. Students will gain experience of protein expression, purification and crystallization, and biochemical assays to investigate oligomerisation of the proteins involved

Supervisor/ host lab
Dr Cara Vaughan, ISMB Lecturer in Structural Biology, Department of Biological Sciences, Birkbeck

This project is suitable for all Biosciences undergraduates

Deadline for enquiries
16 January 2012