[ccp4bb] ccp4mg question
Dear all, I am working with a cyclic protein and I am trying to make a figure with ccp4mg. I would like to know how to say to ccp4mg that the N and C-terminus are bound Although atoms are at a covalent bond distance, the chain is broken by ccp4mg... Many thanks in advance for your suggestions and help! Maria
Re: [ccp4bb] ccp4mg question
Dear all, it seems that none of the tricks from Mark or Tim or Stuart´s ccp4mg making bond strategy works out... In all cases, the bond is made when drawing in think/fat band style... However, when representing in ribbon style, the link is not there... Any other suggestion is welcome! Cheers, Maria Quoting Stuart McNicholas: On 15/11/2012 12:26, SANCHEZ BARRENA, MARIA JOSE wrote: Dear all, I am working with a cyclic protein and I am trying to make a figure with ccp4mg. I would like to know how to say to ccp4mg that the N and C-terminus are bound Although atoms are at a covalent bond distance, the chain is broken by ccp4mg... Many thanks in advance for your suggestions and help! Maria Dear Maria, On e thing to try is to click on the icon next to the molecule name (in the display table). Select Structute definition - Edit bonds. A new window should appear. In the main graphics window right click on one of the atoms you wish to connect and select Add/delete bond - first atom. Then right click on secon atom, select Add/delete bond - (name of first atom). This should draw a bond between them. Best wishes, Stuart McNicholas
[ccp4bb] protein gets trapped
Dear all, I would like to share with you some problems I am experiencing with a protein, in case someone has an idea about what it is going on: I am working with a protein module that gets trapped in Ni-NTA or glutathione beads. If the protein is purified without tag (ion exchange plus gel filtration), the protein can be purified properly. However, when expressing with a His or GST tag, it gets trapped and it cannot be eluted with imidazole/glutathione or even with 8M Urea (in case it is unfolded) Does someone know what can be happening? We do not have any information about an enzimatic activity of the module that might permit the protein bind covalently to the beads Furthermore, we got the crystal structure in complex with another protein and the folding of the module does not resemble anything with enzimatic activity Many thanks in advance for all your suggestions, Maria
[ccp4bb] contaminant when overexpressing a GST tagged protein
Dear all, I am trying to express a eukatiotic protein (E. coli codon optimized sequence) with a GST tag at the N-terminus. I always get my overexpressed protein and a contaminant around 60kDa. This contaminant is not washed out of the column when washing glutathione beads with 1M NaCl-buffer. However, during o/n incubation with proteases that cleave the GST off (thrombin or TEV), the contaminant is in the soluble fraction. Has someone had this experience? I know about contaminants that bind to Ni2+ when overexpressing a His-tagged protein, but this is the first time I get such thing with a GST-tagged protein. One could think that that contaminant could be a protein that binds to my overexpressed protein, but I do not think so, cause I always get a huge band of the contaminant, independently on the amount of the protein of interest Many thanks in advance for all your suggestions and sorry for asking about non-crystallographic topics. Regards, Maria - María José Sánchez-Barrena, PhD Departamento de Cristalografía y Biología Estructural. Instituto de Química Física Rocasolano. CSIC Serrano 119. 28006 Madrid (Spain)
Re: [ccp4bb] XXII IUCr Congress and General Assembly - Madrid (Spain)
Dear Gerard, you are right, the abstract submission has been extended. Best regards, Maria Quoting Gerard Bricogne: Dear Harry, Thank you for the reminder. Upon connecting to the URL given in Martin's e-mail, however, one immediately gets a pop-up saying: New deadline fo abstract submission: April 28 (no doubt to enable everyone to work until the last minute prior to the royal wedding ...). Can you confirm? With best wishes, Gerard. -- On Fri, Apr 15, 2011 at 11:39:14AM +0100, Harry Powell wrote: Hi folks today is the last day to get your abstract in! On 7 Apr 2011, at 14:24, Martin M. Ripoll wrote: Dear colleagues, This is just to remind you that the “XXII Congress and General Assembly” of the IUCr (International Union of Crystallography) will be held in Madrid (Spain) from 22-30 August 2011 and that your Spanish colleagues kindly invite you to participate not only in the most important crystallographic event until 2014, but also to enjoy a fruitful meeting in a sunny and full of life city! All information is to be found in: http://www.iucr2011madrid.es/ although two important dates to remember are: April 15, 2011: Deadline for abstracts submission May 15, 2011: Deadline for Early bird registration All the best and see you in Madrid! On behalf of the Organizing Committee, Martin (Vice-Chairman) Dr. Martin Martinez-Ripoll Research Professor xmar...@iqfr.csic.es Department of Crystallography Structural Biology www.xtal.iqfr.csic.es Telf.: +34 917459550 Consejo Superior de Investigaciones Científicas Spanish National Research Council www.csic.es image003.jpg Harry -- Dr Harry Powell Chairman ECA SIG9 (Crystallographic Computing) Acting Chairman IUCr Commission on Crystallographic Computing http://www.iucr.org/resources/commissions/crystallographic-computing/schools/mieres2011 -- === * * * Gerard Bricogne g...@globalphasing.com * * * * Global Phasing Ltd. * * Sheraton House, Castle Park Tel: +44-(0)1223-353033 * * Cambridge CB3 0AX, UK Fax: +44-(0)1223-366889 * * * ===
