Mario, beside what you were mentioning, I would definitely try a quick soak (10-30 seconds) of the crystals in cryo conditions supplemented with halides such as NaBr, or NaI, at pretty high concentrations (say 0.5 M), then directly freezing without backsoak. If the crystals survive the treatment, with that amount of mols per unit cells and surely some good NCS, you should be able to phase pretty easily. Well, of course SeMet would be the other option... ciao, s
On Sep 30, 2010, at 1:54 PM, Mario Milani wrote: > Dear all, > i have a 30 kDa protein that crystallize so far in three different conditions > but with the same space group. It initially looks like tetragonal (I4, a=141, > b=141, c=208) and then results triclinic (P1, a=141, b=141 c=144, alpha=119, > beta=119, gamma=90), hosting about 24 mol. in the unit cell. Other data: self > rotation shows the presence of 4 peaks with chi=180; molecular replacement > shows the presence of a pseudo-translation peak; DLS made at protein > concentration close to crystal growth conditions shows a Rh compatible with > something like a tetramer with low polydispersity (about 15%). Do you have > any experience with similar ‘asymmetric’ associations? Do you have any > suggestions, beside the addition of ligands to the crystal growth conditions, > in order to get a ‘simpler’ crystallographic assembly? I have some models > (with sequence identity less than 25%) in order to try MR but all trials so > far did not solve the structure (using balbes, molrep, phaser and epmr). Any > suggestion is welcome. > Thank you, > > Mario Milani -- Sebastiano Pasqualato, PhD IFOM-IEO Campus Dipartimento di Oncologia Sperimentale Istituto Europeo di Oncologia via Adamello, 16 20139 - Milano Italy tel +39 02 9437 5094 fax +39 02 9437 5990
