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-----BEGIN PGP SIGNED MESSAGE----- Hash: SHA1 Hi Mark, does your experience also include the geometry of the model? There might be improvements, as e.g. reported from molprobity, because of a better modelling of the temperature factors. I am asking because you mention improvements of the electron density, whereas some improvements in the geometry, like the Ramachandran plot, may not be directly visible in the electron density. Cheers, Tim On 08/09/2013 04:27 PM, Mark J van Raaij wrote: > Omid, how necessary is it that you do TLS refinement? I.e. are you > just doing it to improve your Rs a bit or is the density really > getting better? If the density is not improving, I would just > refine without TLS and avoid any posterior temperature factor > analysis difficulties. While I am sure there are cases where TLS > refinement really helps (cf. published examples), my experience in > our structures is that it does not really make a difference. Also, > in my opinion the TLS model you define must make some physical > sense, i.e. some logical explanation why your structure or domains > of your structure may have internally consistent anisotropic Bs. > Mark > > > > > On 8 Aug 2013, at 23:01, Ethan Merritt wrote: > >> On Thursday, August 08, 2013 01:51:34 pm Omid Haji-Ghassemi >> wrote: >>> Dear Robbie, Marcus and Reginald, >>> >>> Thanks again for your replies, I truly appreciate the help. >>> >>> The B-factors was set to 20 when performing TLS refinement so I >>> don't think that is the problem. >>> >>> I also tried Marcus's suggestion using output from coot, with >>> no luck. >>> >>> The only thing left to try is to test alternative TLS group as >>> Reginald have suggested. >> >> You have only told us about an increase in average B, not whether >> it is uniformly inflated. Possibly the output from analysis by >> the Parvati server http://skuld.bmsc.washington.edu/parvati would >> indicate specific parts of your structure that are behaving badly >> during refinement. >> >> Ethan >> >>> >>> Cheers Omid >>> >>>> Hi Omid, >>>> >>>> Sometimes the choice of TLS groups and to a lesser extent the >>>> initial B-factor matter a lot. You should try a few other TLS >>>> group selections and see if these give nicer results. Things >>>> to try: TLSMD, including or excluding ligands and >>>> carbohydrates, other common-sense or gut-feeling structure >>>> partitionings. If you have a lot of different groupings to >>>> test, you can reset the B-factor and do pure TLS refinement >>>> (i.e. 0 cycles of restrained refinement) for all of them. You >>>> can then use the best one for your 'final' refinement. It's >>>> much faster then trying your final refinement with all TLS >>>> groups selections. >>>> >>>> Cheers, Robbie >>>> >>>> Sent from my Windows Phone ________________________________ >>>> Van: Omid Haji-Ghassemi Verzonden: 8-8-2013 21:55 Aan: >>>> [log in to unmask] Onderwerp: Re: [ccp4bb] TLS refinement >>>> and ANISOU records >>>> >>>> Dear Ethan, >>>> >>>> Thank you for your reply. >>>> >>>> I will try to review my refinement protocol once more; >>>> however, I am still perplexed at what lies at the heart of >>>> the problem. >>>> >>>> Overestimation of average B-factor using TLS is perfectly >>>> sound, but I am not sure why all my structures the average >>>> increases tremendously. >>>> >>>> In one case it increases from 16.36 to 73.02 for a 2.3Ang >>>> structure. >>>> >>>> I already tried changing weights and number of TLS rounds, >>>> which resulting in only a small change in average B. >>>> >>>> Omid >>>> >>>>> On Thursday, August 08, 2013 11:39:22 am Omid Haji-Ghassemi >>>>> wrote: >>>>>> Dear all, >>>>>> >>>>>> I was about to deposit a few structures to the pdb when I >>>>>> noticed the mean B-factors were larger than one might >>>>>> expect. >>>>>> >>>>>> All the structures were refined using TLS refinement. >>>>>> >>>>>> During refinement in Refmac the average temperature >>>>>> factors for each structure is reasonable. For example, a >>>>>> structure at 2.75� has a mean B-factor of 40; however, >>>>>> after adding the ANISOU records as required by the PDB, I >>>>>> noticed the average B-factors double. >>>>> >>>>> Please see my paper: E. A. Merritt (2011). "Some Beq are >>>>> more equivalent than others". Acta Cryst. A67, 512-516. >>>>> <http://skuld.bmsc.washington.edu/parvati/ActaA_67_512.pdf> >>>>> >>>>> >>>>> In short, the quantity stored in the "B" field of a PDB file after TLS >>>>> refinement is Beq, which overestimates what the isotropic B >>>>> factor would have been if you had refined without TLS. So >>>>> in general the "average B" after TLS refinement is always >>>>> higher than the "average B" without TLS. The problem is >>>>> that the two quantities marked "average B" are not directly >>>>> comparable. >>>>> >>>>> Having said that, the overestimate is not usually as much >>>>> as a factor of 2. So something else may indeed be causing a >>>>> problem in your case. >>>>> >>>>> Ethan >>>>> >>>>> >>>>>> >>>>>> Is this normal? >>>>>> >>>>>> Sincerely, Omid >>>>>> >>>>>> --------------------------------------------------- >>>>>> --------------------------------------------------- Omid >>>>>> Haji-Ghassemi, Graduate Student Department of >>>>>> Biochemistry & Microbiology University of Victoria PO Box >>>>>> 3055 STN CSC Victoria, BC, V8W 3P6 CANADA >>>>>> >>>>>> Tel: 250-721-8945 Fax: 250-721-8855 >>>>>> >>>>> >>>>> -- Ethan A Merritt Biomolecular Structure Center, K-428 >>>>> Health Sciences Bldg University of Washington, Seattle >>>>> 98195-7742 >>>>> >>>> >>> >> >> -- Ethan A Merritt Biomolecular Structure Center, K-428 Health >> Sciences Bldg University of Washington, Seattle 98195-7742 > - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -----BEGIN PGP SIGNATURE----- Version: GnuPG v1.4.14 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFSBP3LUxlJ7aRr7hoRAo2eAKDoDFsAnywjd3GIc2S+d7UZw1PU2QCffIn4 UZiFSH1yXDMexQQIkzyBSqI= =Y4ad -----END PGP SIGNATURE-----