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List of Projects for TSMB '08-'09

Note: Make sure you read the paragraph describing each project carefully before making your choice. Students who produce their own clear, relevant diagrams, using appropriate software, will be marked significantly higher than those who copy equally relevant diagrams from elsewhere with due acknowledgement.

Fragment based Drug Design
The use of small chemical fragments that bind with low (mM) affinity to drug targets as starting models for drug design is a relatively new development. Biophysical (NMR, SPR, Calorimetry, X-ray Crystallography) are used to screen libraries of small molecules. From a weakly binding compound a series of larger compounds are designed to find a molecule with tighter binding (nM) suitable for use as a drug. Although the affinity of these small molecules is low, the ligand efficiency (affinity/molecular weight) is often very good. Amongst the companies that developed this method are Astex pharmaceuticals (X-ray) and Abbot Laboratories (SAR-by-NMR). The project will describe these approaches and some of the successes.

Electron Microscopy, Crystallography and Molecular Modelling
Detailed molecular structures of many large protein complexes or "molecular machines" have been produced using a combination of techniques. This is done by fitting atomic resolution structures of individual protein chains (or even domains) into the lower-resolution electron density maps from electron microscopy (or from low resolution X-ray crystallography). Describe the methods for fitting structures determined by X-ray crystallography or NMR into low resolution maps and give examples of how this has been used in practice, what biological insight it has given, and the limitations that arise from the low resolution.

Methods for Automated Crystallographic Model Building
Increasingly automated methods can be used for building a nearly complete atomic model (PDB) file from an initial experimentally phased electron density map or molecular replacement solution. These programs include ARP/wARP, Bucanner (CCP4), Resolve and Textal (Phenix). The project will outline how the various automated building programs work and describe what applications the methods are effective for and their limitiations.

Structural Studies of the Kinetochore
The kinetochore is a macromolecular assembly of proteins that bridges the sister chromatids to the microtubules of the mitotic spindle. Correct formation and assembly of the kinetochore is essential for the faithful segregation of sister chromatids during mitosis, a process that is essential for cell division. Errors in this process lead to aneuploidy - uneven segregation of chromosomes between the mother and daughter cell. This leads to genetic instability, a common phenotype of certain cancers. The aim of this project is to survey the literature of crystal and EM structures of the proteins of the kinetochore, both mammalian and yeast. This should also include checkpoint proteins involved in the signalling for anaphase wait. Reference should be made to the methods used in the structure determinations and why these methods were applied in the particular cases.

Structural Studies of Amyloid plaques and Fibrils
Alzheimers disease and a number of other brain degenerative diseases are associated with the build up of plaques and fibrils in the brain. Describe how different structural biology techniques are being used to study these characteristic structures and the molecules that make them up. How has structural information helped our understanding of the mechanism of pathogenesis? In addition describe how structural biology is being used in the design and development of novel, mechanistically based, drugs against these devastating conditions.

Solid state NMR of proteins
Solid state NMR allows the study of larger protein complexes and membrane proteins than traditional solution protein NMR. Describe how structure determination by solid state NMR differs from solution state methods. Give examples of protein structures that have been determined by solid state NMR commenting on their quality, and their scientific impact.

Measuring progress in protein structure prediction: the CASP series of structure prediction competitions.
CASP (Critical Assessment of Structure Prediction) http://predictioncenter.org/ is a series of biannual "competitions" to assess the quality of protein structure prediction methods. The results of each competition are published in a supplement to the journal Proteins and there are reviews published after each CASP conference. This project will involve reviewing the progress made in structure prediction during the first eight competitions (from 1994 to 2008), and critically discussing the most successful approaches and methodologies.

Phosphoinositide binding domains in the PDB
Phosphoinositides are one of the most important classes of cell signalling molecules. There are several types of protein domain that bind them, some specific for certain inositides while others bind inositides nonspecifically. The project should use the Protein Data Bank and the literature to describe the structures of these domains and how discrimination between different inositides is achieved.

Prevention and use of radiation damage in protein crystals
Radiation damage is a major problem with protein crystals particularly at third generation synchrotrons. The project will present an overview of the causes of radiation damage, its prevention and how it can be used to the experimenter's advantage. Work particularly by Elspeth Garman has tried to systematically understand the problem and find methods of prevention. Raimond Ravielli and colleagues have also deliberately used radiation damage to derive phase information and to study which part of the protein is damaged first.

Restriction/methylation systems and their structure and use in molecular biology
This project requires analysis of the scientific literature and on-line resources to discuss the various types of restriction/methylation systems. The aim is to give an overview of their biological role, their use in DNA manipulation methods and to discuss the relationship of structure to function for at least one example of a methylase and one endonuclease within the overall context of what is known about these enzymes from a structural perspective.

Structural Studies of Filaments
Filaments can be studied by electron microscopy and X-ray fibre diffraction. Describe these methods and discuss the biological insights that have come from them. The most notable examples are from the cytoskeleton. There are examples of both methods in section 12 which could form a starting point for this project. You should look to give examples of at least three applications of each method to different proteins.

Crystallisation Screens and Optimisation.
There are a number of companies who produce commercial screens to find initial conditions for the crystallisation of macromolecules. These include Hampton Research, Molecular Dimensions Ltd, NEXTAL (part of Qiagen), Jena Bioscience and Emerald. Using material from the companies web sites and the academic literature on crystallisation, outline the underlying science these screens are based on - how the screens were designed, what types of system are they used for, and consider what are the strengths and limitations of the approach. The project should end by describing how to progress from initial small crystals (obtained from a commercial screen) to single crystals suitable for X-ray diffraction.

SAXS/SANS (Small angle X-ray and neutron scattering)
Small angle X-ray and neutron scattering can be used to probe the low resolution shape, conformation and oligomeric state of macromolecules. In some cases to a similar resolution as electron microscopy. The project will outline the theory of the methods and concentrate on on at least four detailed examples where the results have given biological insight.

Tomography by Electron Microscopy
Sectioning of vitrified cell and tissue samples has been pioneered by J Dubochet and tomography of vitreous samples by W Baumeister. Tomography of vitrified sections provides a view of cells and tissues in their native, hydrated state, without the severe artefacts of fixation, embedding in plastic resin and thin sectioning. Write a review of these important methodological developments and their significance for cell and molecular biology.

A Survey of the Precision of PDB Data
Protein structures deposited in the PDB vary a great deal in quality. Some structures are only available at low resolution and a few, still, may even be just plain wrong! You will conduct a survey of the 37,000+ structures in the PDB looking for trends in the usually reported measures of structural accuracy and precision, including resolution and R-factor. How do resolution and R-factor vary with protein size and type, and from the 1970s to the 2000s?

A Comparison of BLAST and FastA as Sequence Analysis Tools
BLAST and FastA are the programs of choice for the common bioinformatics task of searching a protein database for sequences that are homologous to a test sequence. Explain in detail how each of these programs works, and compare and contrast them in terms of speed, precision, and accuracy. Include the protein specific tools PSI-BLAST and PHI-BLAST in your discussion of BLAST.

FRET (Fluorescence Energy Resonance Transfer) and FLAP (Fluorescence Localisation after Photo Bleaching) in vivo
Fluorescence techniques such as FRET and FLAP are increasingly being used to study protein interaction and mobility in intact cells. Describe these methods, giving examples of the type of studies that can be carried out with the techniques and their limitations.

Synchrotron Radiation Circular Dichroism
Synchrotron radiation can be used to extend the wavelength range and the type of samples on which Circular Dichroism data can be collected. Prof. Bonnie Wallace in our department is one of the pioneer's in this field and starting by searching the literature with her name will give a good starting point for your research. The project should describe SRCD and discuss its advantages and the biological results it has produced so far giving at least four detailed examples.

Please refer to the guidelines before choosing your project.

Nicholas Keep and Clare Sansom, June 2009