The convention for "P22121"s is P21212, which is used by both CNS and CCP4 and many else, if not all. The unit cell needs to be reindexed (a-b-c --> b-c-a in your case). Then please try again. Lijun
> Thank you for your suggestions. > 1. The unit cell of my crystal is 47.41 99.67 114.97 90 90 90 space group > P22121, different from PDB structure 64 64 113 90 90 90 P43212, which has > the same growth condition. > 2. Mathhews_coef indicate my crystal should be dimer if ~30kD. I used all > monomer, dimer and tetramer PDB templates and their truncated models, but > all high R-factor. > 3. My protein has only one domain(ligand binding domain), and there are > both > structures reported with or without ligand. > > I think even though my protein was degraded, the structure can be > determined > due to its remained same sequence. > So now do I have to turn to mass spec? > > 2008/7/8 Zhijie Li <[EMAIL PROTECTED]>: > >> Hi Haitao, >> >> I need to ask you a few questions first: >> >> 1. Did you mean you could not solve your structure by molecular >> replacement? Did you compare your crystal's unit cell with the PDB file? >> Are >> they significantly different? If the assymetric unit has more than one >> monomer, have you tried doing a molecular replacement search with one >> monomer only? >> >> 2. Can you give us the PDB number so that we can take a look at the >> protein? The reason for that is, I suspect that your protein was not >> degraded from either end, but only got clipped some where on the surface >> - >> so the structure is basically unperturbed. >> >> If the PDB structure turns out to be single-domain, then you should do >> your >> molecular replacement search with the whole protein. If it is a >> two-domain >> structure, and one of them is ~20kD, then try use that 20kD domain to do >> the >> search again. >> >> R-fac~=0.5 is probably saying that your current solution is totally >> wrong. >> As I remember, R-factor for a totally radom acentric (for example, >> protein) >> structure is 0.59. >> >> Also, Even if you follow the published crystallization conditions, your >> protein may still crystallize in a totally different way. But unless the >> protein itself has changed its shape (which normally does not happen), >> you >> should be able to do a molecular replacement. >> >> If molecular replacement does not work at all, then maybe it is time >> to send your sample to mass spec to see what it really is. But I highly >> suspect what you need to do now is nothing but to optimize your >> molecular >> replacement parameters. >> >> Zhijie Li >> Graduate student, Univeristy of Toronto >> >> >> >> ----- Original Message ----- >> >> *From:* Haitao ZHANG <[EMAIL PROTECTED]> >> *To:* [email protected] >> *Sent:* Monday, July 07, 2008 10:27 PM >> *Subject:* [ccp4bb] Truncated protein structure >> >> Dear all, >> I repeated a protein crystallization which is reported in PDB with the >> almost same condition, and got a 2.6A data. But the problem is that I >> can >> not determine the right phases with neither CCP4 nor CNS. >> Then I found the protein in my crystal had been degraded from ~30kD to >> ~20kD by SDS-PAGE, but did not know from which terminus it was >> truncated. >> So I used truncated PDB templates which N-, C- or both terminal were cut >> to >> fit the length of my shorter protein, and tried many different >> templates. >> But always high R-factor (~0.5). >> With COOT I found the backbone did not fill the electron density map >> perfectly. >> The only difference of my crystallization condition is 277K(mine) >> *vs*295K(reported) in temprature. >> >> Any suggestions will be appreciated. >> >> >> Haitao ZHANG, Ph.D Student, >> Shanghai Institute of Materia Medica, >> Chinese Academy of Sciences >> >> >
