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Faculty of Biological Sciences School of School of Molecular and Cellular Biology Summer Research Placement 8 weeks, between July – September 2016 Job Summary The Picornaviridae family of viruses contain many important pathogens, such as poliovirus and hepatitis A virus which cause devastating diseases in mammals. Members of the Picornavirales have a single stranded, positive sense RNA genome encapsidated in a non-enveloped capsid with pseudo T=3 icosahedral symmetry. Despite detailed structural information for the capsid of Picornavirales, the mechanism for genome encapsidation is not understood. Cowpea mosaic virus (CPMV) is a plant-infecting member of the Picornavirales which is of major interest in biotechnology, and is an ideal model system to study genome encapsidation. A better understanding of CPMV genome encapsidation will allow us to further develop CPMV biotechnology applications. We aim to use high-resolution cryo-electron microscopy (EM) analysis of CPMV and CPMV mutants to unravel the mechanism of genome packaging. We have produced an atomic resolution cryo-EM structure of wild-type (WT) CPMV and an empty virus-like (eVLP). Using the information from both eVLP and CPMV maps we have designed, produced and purified mutant CPMV particles that are unable to package RNA. A F192W mutant, contains a conservative mutation of an amino acid located on the external surface of CPMV, which is unable to encapsidate RNA but can produce viral particles. You will analyse the structure of CPMV F192W using cryo-EM. You will make and assist in screening cryo-EM grids of CPMV F192W to ensure high quality grids are produced using our current 200 kV electron microscope. Optimal grids will be analysed using our new 300 kV Titan Krios electron microscope (due to be delivered May 2016). We will collect a large cryo-EM data set for the CPMV F192W mutant. We anticipate collecting ~1,500 micrographs using a direct electron detector. We aim to produce high-resolution reconstructions of the CPMV mutant using single particle image processing software (RELION) to produce a 3D map. Using this method we have already produced high-resolution WT CPMV and an CPMV empty virus like particles (eVLP) maps (3.4 Å and 3.0 Å respectively) (Hesketh et al., Nat. Comms, 2015). We will compare the structure of WT CPMV to the structure of the mutant CPMV to gain further insights into CPMV biogenesis. Using this information we aim to understand better how CPMV recognises it’s RNA genome for encapsidation prior to capsid assembly. Stipend is £200 per week. Informal enquiries may be made to Emma Hesketh, email: [email protected]. To apply, please send your CV and a covering letter (200-300 words maximum) Closing Date: Noon on 9th March 2016 Interviews: 16th March Job Description Reports to: Emma Hesketh Main Duties and Responsibilities Prepare samples for negative stain and cryo-electron microscopy (cryo-EM). Process cryo-EM data using single particle image processing software such as RELION. Write up technical procedures and methods into detailed protocols, to allow reproducibility of experiments by others. Present data in an understandable manner to members of the group. Person Specification Essential Ability to work independently after induction period, manage and prioritise own tasks on a day-to-day basis with little supervision. Good organisational skills and time management skills. Highly motivated and hard working. Desirable Knowledge of UNIX and the command line. Additional Information Details of the terms and conditions of employment for all staff at the university, including information on pensions and benefits, are available on the Human Resources web pages accessible via the links on the right hand side, or at http://www.leeds.ac.uk/hr/index.htm Criminal Record Disclosures A Criminal Records Disclosure is not required for this position. However, applicants who have unspent convictions must indicate this in the ‘other personal details’ section of the application. Disabled Applicants The post is located in the Faculty of Biological Sciences. Disabled applicants wishing to review access to the building are invited to contact the department direct. Additional information may be sought from the Recruitment Officer, email [email protected] or tel + 44 (0)113 343 1723. Disabled applicants are not obliged to inform employers of their disability but will still be covered by the Disability Discrimination Act once their disability becomes known. Further information for applicants with disabilities, impairments or health conditions is available in the applicant guidance. University Values All staff are expected to operate in line with the university’s values and standards, which work as an integral part of our strategy and set out the principles of how we work together. More information about the university’s strategy and values is available at http://www.leeds.ac.uk/comms/strategy/