Hi, Lattice-translocation disorder is one possibility. How does the diffraction image look like? Are there any streaks?
Best regards, Takanori Nakane On 2014-07-30 15:58, Minyun Zhou wrote:
Dear all, I am trying to determine the structure of a protein-DNA complex. I collected several datasets of the same protein in complex with two dsDNAs. Two DNAs have the same sequence, but one DNA contain a single central mispair, which is cleaved by the enzyme leaving an abasic site, while the other contains a non-cleavable mispair. Two datasets have almost the same space group and cell parameter, however, the later one has been detected with pseudo-translation while the other not. The details of two datasets are as follows: Dataset 1. Protein with abasic site containing dsDNA: C2221: a=106, b=111, c=182.2, α=β=γ=90 Resolution: 2.45 A Xtriage summary: The largest off-origin peak in the Patterson function is 18.57% of the height of the origin peak. No significant pseudotranslation is detected. Dataset 2. Protein with non-cleavable lesion containing dsDNA: C2221: a=106.1, b=111.1, c=183, α=β=γ=90 Resolution: 3.0 A Xtriage summary: The analysis of the Patterson function reveals a significant off-origin peak that is 60.75 % of the origin peak, indicating pseudo translational symmetry. I first determined the structure using the dataset 1 by MR. There are two protein-DNA complexes in one asymmetric unit, and two molecules are similar except the conformation of a small loop at the active site. The final model has the Rwork/Rfree of 19.3%/23.0%. Then I use this refined structure as model to solve the second structure with dataset 2. The final Rwork/Rfree is 24.3%/28.9%. The DNA density looks okay while the protein part is problematic. After refinement, although the main chain of the protein is fitted well into the density, there is still a lot of unidentified continuous positive density next to the polypeptide chain, especially near the region involved in the crystal packing. I attached a snapshot below, which shows one of these positive densities lies along a helix. Since there is no other polymer in the reservoir solution that can fit, the additional density seems to indicate another slightly shifted conformation of the protein itself. My question is whether this seemingly “dual conformation” is caused by pseudo-translation and indicates the solution I found is incorrect. And how can I eliminate the effect of pseudo-translation to get the correct structure? Thanks for your help! Best regards, Minyun
