Re: [ccp4bb] process data in R32

As far as I know all the programs we use will handle the non-primitive hexagonal setting of rhombohedral space groups (ie a=b != c, alpha=beta=90, gamma =120), often denoted "H3" or "H32" (aka R3:H in cctbx). Many of them will probably handle the primitive rhombohedral setting (a=b=c, alpha=be

Re: [ccp4bb] advice for processing data from multiple small wedges

Hi Tony, in my experience it is not uncommon for crystals that diffract to resolution worse than 3 A to be anisomorphous - after all, there is a reason for diffracting badly, and that must be the lattice defects. Lattice defects e.g. often lead to differences in cell parameters, and in that case

[ccp4bb] PEG in the pdb?

Hi everybody, I just solved the structures of an enzyme an some variants. In the active site cavity of each variant I found one or two fragments of PEG1000 bound. I used PEG1000 in the crystallization condition. Among the enzyme variants the number of non-hydrogen atoms of these PEG fragments v

Re: [ccp4bb] PEG in the pdb?

Dear Klaus depends how many missing residue you want advertised in the header of your deposited file. The pdb will put in a special Remark 610 and Remark 615 to indicate any missing residues from your heterogen list. However the policy is not to list every missing (or zero occup

[ccp4bb] Protein Data Bank in Europe roadshows and tutorials

The Protein Data Bank in Europe (PDBe; http://pdbe.org/) offers a number of educational resources as part of its wider mandate for outreach and training within the European Bioinformatics Institute (EBI; http://www.ebi.ac.uk/). As part of this commitment, we offer the PDBe "roadshow" (http://p

Re: [ccp4bb] process data in R32

Coming from small molecule crystallography where this problem simply doesn't exit, I found it very difficult to understand all the fuss! The small molecule crystallographers specify the symmetry operations rather than the space group so there is nothing special about using different settings. If a

Re: [ccp4bb] process data in R32

Hi Ray, In HKL2000, by default (i.e. - without changing anything manually) for R32/H32, Denzo/Scalepack leads to a .sca file that has hexagonal unit cell parameters in the header but has the space group listed as R32. Scalepack2mtz in ccp4 does not like this mismatch so you have to simply cha

[ccp4bb] Unidentified density.

Dear All, I am refining structure of protein at 1.7A. It is enzyme with 3 histidine residues in the active site, which are chelating metal (at the moment I built in calcium but I do not know for sure, which metal is bound there). I can see additional density "on top" of metal, which I can

Re: [ccp4bb] calculating solvent volume from molecular surfaces

Take a look at MAMA from Uppsala Software Factory http://xray.bmc.uu.se/usf/mama_man.html It can generate the mask from the PDB file. This will, however, leave internal cavities belonging to "solvent". If you don't want that, the following MAMA script will fix it giving you the mask of the pro

Re: [ccp4bb] PEG in the pdb?

PEG solutions contain fragments of all sizes - it is the average size (however defined by the manufacturer) that is 1000. So technically it is incorrect to claim that you have PEG1000 molecules bound to your protein, it is most likely much shorter fragments that can penetrate the channels in prote

Re: [ccp4bb] PEG in the pdb?

On Thu, 2010-08-12 at 08:57 +, MARTYN SYMMONS wrote: > Zero occupancy is generally a deprecated way of dealing with missing > density as it is confusing for less experienced user of the > coordinates. I think zero occupancy can be useful during refinement as > the atoms help fill space (or for

Re: [ccp4bb] Unidentified density.

Nevertheless, what do you have in mother liquor/protein buffer? On Thu, 2010-08-12 at 17:24 +0200, wrote: > Dear All, > > I am refining structure of protein at 1.7A. It is enzyme with 3 > histidine residues in the active site, which are chelating metal (at > the moment I built in calcium but

[ccp4bb] Off topic: PKa of protein-DNA complex.

Hi, Is there any server or program can calculate the theoretic PKa of the protein-DNA complex? Given the structure of the protein and the sequence of the DNA. Any suggestion will be appreciated! Best Yang

Re: [ccp4bb] Unidentified density.

Could this be a metal cluster? Is your protein solution colored at all? Have you made an anomalous difference map? Is there reductant in the mix? What small molecules were in the mother liquor? JPK - Original Message - From: "" To: Sent: Thursday, August 12, 2010 10:24 AM Subject:

[ccp4bb] Suggestions for Reducing Protein Precipitation

Hi. I am working with a protein that has difficulties staying in solution when concentrated. The buffers that I have been using contain 20 mM Tris pH 8.0, 5 mM BME (or 2 mM DTT), 10% Glycerol, and NaCl from 50 mM up to 2 M. The protein seems fine to dialyze into low salt buffer (50 mM NaCl) when

Re: [ccp4bb] Suggestions for Reducing Protein Precipitation

Hi Matt, Check out the following paper and some screens available commercially based on these: Acta Crystallogr D Biol Crystallogr. 2004 Sep;60(Pt 9):1670-3. Epub 2004 Aug 26. Optimum solubility (OS) screening: an efficient method to optimize buffer conditions for homogeneity and crystallization

Re: [ccp4bb] Suggestions for Reducing Protein Precipitation

Matt, I have found the optimum solubility screen to be helpful: Optimum solubility (OS) screening: an efficient method to optimize buffer conditions for homogeneity and crystallization of proteins. Jancarik J, Pufan R, Hong C,

Re: [ccp4bb] Suggestions for Reducing Protein Precipitation

Hi Matt, Have you tried changing the pH? Is it possible that at pH 8 you are at the pI of your protein (i.e. where it has zero net charge and is at its least soluble) ? I have also read a paper (can't find the reference right now) where the authors precipitated their protein by dialysing into wat

Re: [ccp4bb] PEG in the pdb?

That's a good point, Ed Based on the formula: HO CH2-(CH2-O-CH2)n-CH2OH, PEG MW = 44n+62, a table of n to MW goes like below which gives an idea of what range is possible. Someone maybe knows what polydispersity can be expected from the synthetic process. As you say though, a specific range cou

[ccp4bb] heavy atom soak

Dear All, In order to phase, I intend to derivatize my protein(30kDa)-DNA(7.2kDa) complex with heavy atoms. I wanted to know which was the better way to do it: longer soaks at lower heavy atom concentration, or shorter soaks with higher concentration of heavy atom. Also, what concentrations and t

Re: [ccp4bb] PEG in the pdb?

Googling peg polydispersity returns this as the second hit http://www.springerlink.com/content/tp643l7678048447/ which is circa 1973 data from behind the iron curtain. The technology may have improved, and the most recent I can quickly find is this paper Plata et al., Electrophoresis, (2010), 3

Re: [ccp4bb] Suggestions for Reducing Protein Precipitation

Hi all. I found that reference. :0) Acta Crystallogr D Biol Crystallogr. 2006 Jul;62(Pt 7):833-42. Epub 2006 Jun 20. Assessment of a preliminary solubility screen to improve crystallization trials: uncoupling crystal condition searches. Izaac A, Schall CA, Mueser TC. http://www.ncbi.nlm.nih.gov/

Re: [ccp4bb] heavy atom soak

Hi Amit, For heavy atom phasing, you'll have to try all the things you can, limited only by supply of crystals, availability of heavy atoms (and considering heavy atom handling and waste disposal policies for solutions, tips, etc.) and time and effort. I had success with both short soaks (tri