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Re: [ccp4bb] process SeMet labelled data |
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CCP4bb navigationCCP4bb <-- 2007 <-- March 2007 <-- 01 March 2007Subject: Re: process SeMet labelled data From: James {- dot -} Whisstock {- at -} MED {- dot -} MONASH {- dot -} EDU {- dot -} AU Date: 2007-03-01 Below I've tried to give you some broad ideas about where to look / read and some packages / approaches to try. For a start there is a very comprehensive tutorial on the ccp4 site together with the experiemental phasing roadmaps. http://www.ccp4.ac.uk/dist/examples/tutorial/html/heavy-tutorial-mad.html which goes through the process step by step. For indexing and processing images, you are probably using MOSFLM or HKL2000?, both of which have excellent manuals that help with troubleshooting. At this stage you need to pick the appropriate bravais lattice then you need to refine and integrate. Once you have integrated (run in ccp4 for MOSFLM), then you need to scale the data using, for example, SCALA. Have a look at the following tutorial for running scala. http://www.ccp4.ac.uk/courses/ECM2004/runningscala.pdf Don't forget to seperate anomolous pairs when running scala!!!! Here, if relevant for your bravais lattice, you can also start to look at systematic absences to try to assign / narrow down the options for your space group. Remember, if your space group is incorrect, then (even though you may have a derivative) you won't be able to solve your structure. Thus if there are ambiguities, you need to systemically try out the options. Once you have scaled your data you may merge etc (see tutorial) using CAD and SCALEIT and start looking at some Anomalous Difference Patterson Maps to identify Heavy atom sites. see http://www.doe-mbi.ucla.edu/~sawaya/tutorials/Phasing/anopat.html Also, have a look at using SHARP/autoSHARP http://www.globalphasing.com/sharp/manual/index.html or SOLVE/RESOLVE http://www.solve.lanl.gov/ to analyse your data. These are both excellent packages for processing MAD / SAD datasets and calculating heavy atom positions / phases. For SHARP / autoSHARP you can supply the individual unmerged mtz's. SHARP will also alert you in a good humoured fashion to any deficiencies in your data and will try to pick the best approach (for e.g. it may try SAD depending on your data quality etc). autoSHARP will also run solvent flattening, give you final flattened maps readable in coot (which you should inspect to see if you can see features such as helices / strands etc). autoSHARP will even input your data into ARP/wARP, and if you are really fortunate you might even come in in the morning (watching ARP chain tracing can be rather addictive so its really best to go home!) and find that ARP has done a good job at a preliminary model or give you a nice starting point for manual building! Also for lower resolution and where you can clearly see features in the map for preliminary chain tracing try using Bucanneer. Hope this helps, good luck and above all have fun! Cheers James Dirk Kostrewa > Hi Mark, > > although Shivesh's question was not very specific, and he should have > clearly given some more informations about what he would like to know, > he is probably a beginner in crystallography and simply asked for help > on this board. Not everyone has always time or is always in the mood to > answer such questions. In my opinion, it's then better not to respond at > all than to give an answer like yours that is neither helpful nor funny! > We all should try to keep a good style here. > > Dirk. > > Mark J. van Raaij wrote: >> why don't you just send all your images to the ccp4bb, then we'll >> process them, solve the structure and publish it for you. >> And we might put you in the acknowledgements, if you are lucky. >> Mark >> On 28 Feb 2007, at 16:35, Jonathan Grimes wrote: >> >>> Anastassis Perrakis wrote: >>>> On Feb 28, 2007, at 14:37, shivesh kumar wrote: >>>> >>>>> Dear all, >>>>> I have a data set at 2.2A, of the selenomethionene labelled >>>>> protein.How should I process the data. >>>> >>>> Carefully ! >>>> >>>>> Thanx for the help. >>>>> Shivesh >>>> >>>> Tassos >>> >>> >>> i am sure what tassos really meant was "Very Carefully !" >>> >>> jon >>> >>> -- >>> Dr. Jonathan M. Grimes, Royal Society Research Fellow University >>> Research Lecturer >>> Division of Structural Biology >>> Wellcome Trust Centre for Human Genetics >>> University of Oxford >>> Roosevelt Drive, >>> Oxford OX3 7BN, UK >>> >>> Email: Jonathan@strubi.ox.ac.uk, Web: www.strubi.ox.ac.uk Tel: >>> (+44) - >>> 1865 - 287561, FAX: (+44) - 1865 - 287547 >> >> Mark J. van Raaij >> Dpto de Bioquímica, Facultad de Farmacia >> and >> Unidad de Rayos X, Edificio CACTUS >> Universidad de Santiago >> 15782 Santiago de Compostela >> Spain >> http://web.usc.es/~vanraaij/ >> >> >> > > > -- > > **************************************** > Dirk Kostrewa > Paul Scherrer Institut > Biomolecular Research, OFLC/110 > CH-5232 Villigen PSI, Switzerland > Phone: +41-56-310-4722 > Fax: +41-56-310-5288 > E-mail: dirk.kostrewa@psi.ch > http://sb.web.psi.ch > **************************************** -- Professor James Whisstock NHMRC Principal Research Fellow / Monash University Senior Logan fellow Department of Biochemistry and Molecular Biology Monash University, Clayton Campus, PO Box 13d, VIC, 3800, Australia +613 9905 3747 (Phone) +613 9905 4699 (Fax) +61 418 170 585 (Mobile) CCP4bb navigationCCP4bb <-- 2007 <-- March 2007 <-- 01 March 2007 |
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