[ccp4bb] Comparison of membrane protein expression
Dear all I want to try a factorial approach for my His-tagged membrane proteins. The screening parameters are E. coli strains, temperature and media. Is there any simple but representative method I can use to compare the expression level without isolating the membrane? Is whole cell lysis and Western blot a good method? Thank you. Theresa
Re: [ccp4bb] Comparison of membrane protein expression
Hi, You can have a look at the paper titled The funnel approach to the pre-crystallization production of membrane proteins by Oded Lewinson, Allen T. Lee, and Douglas C. Rees. They have done high throughput over-expression of various membrane proteins. -- Nazia Nasir PhD Scholar Protein Crystallography Lab National Institute of Immunology New Delhi On Sun, Apr 28, 2013 at 10:56 PM, Theresa Hsu theresah...@live.com wrote: Dear all I want to try a factorial approach for my His-tagged membrane proteins. The screening parameters are E. coli strains, temperature and media. Is there any simple but representative method I can use to compare the expression level without isolating the membrane? Is whole cell lysis and Western blot a good method? Thank you. Theresa
[ccp4bb] Poor electron density in some of the chains in an asymmetric unit
Dear All, We are working on a crystal structure with 12 molecules in an asymmetric unit. It is a heteromer and each chain is made up of two type of chains. Therefore there are 24 chains in an asymmetric unit. It is giving solution in P1. Rfactor and Rfree has reached 20.25 and 25.6 at a resolution of 1.8Angstrom All the chains are refining well and have good electron density except 3 chains which have very poor electron density in most of the region. Whereas, the crystal structure of variants of the same protein we have solved earlier refined well and has very good electron density in all the chains. What could be the possible reason for these three chains which are not refining properly. Any suggestions will be very useful. Thanks and Regards -- Sonali Dhindwal “Live as if you were to die tomorrow. Learn as if you were to live forever.”
[ccp4bb] PhD and Postdoctoral positions in structural virology at the Duke-NUS GMS
A PhD student and a postdoctoral position are available at The Duke-NUS GMS (Singapore) under the supervision of Dr Shee-Mei Lok. In our laboratory, we use a combination of techniques such as x-ray crystallography and cryo electron microscopy to investigate dengue virus complexes and its structural changeduring viral infection.We have the state-of-the-art facilities in cryoEM such as The FEI Titan Krios, phase plate, liquid cell system to investigate both frozen and liquid forms of the sample. For the postdoctoral position, candidates with a Ph.D in structural biology such as x-ray crystallography and cryoEM within 2-3 years of their graduation are encouraged to apply. Interested candidates are requested to send their CV, summary of research interests and names of 3 references to sheemei@duke-nus.edu.sg. Regards, Shee-Mei Lok Assistant Professor Program in Emerging Infectious Diseases, Duke-NUS Graduate Medical School, KTP Building Level 9 Room 19 8 college Road, Singapore 169857 Ph: 65-65165840 Fax:65-62212529
