Hi,<br><br>      I have a 3.2A dataset for a protein-DNA complex. The protein is a homodimer, and the DNA is almost palindromic (except one base pair in the middle and two or three base pairs at both two ends). It is my first time solving structures, and unfortunately the resolution is low. No body in our lab has used ccp4 or phenix, so I am really frustrated as a second year student. <br>
      I mainly used ccp4. So far, the best R/Rfree I got is 0.27/0.34.  I went to the crystallography meeting, and people suggested me to rely more on geometry. I remember I got a DNA restraints file and a refmac script from someone on this mailing list, and that really helped (otherwise the DNA base pairing will be weird). Can someone tell me how to restraint the protein (helix)?  I can only see a few side chains in the helix, so it&#39;s hard to say whether the registration of the helix is correct or not. Maybe my high R-free is due to the helix in the wrong position?<br>
      People also suggested me to include NCS and TLS in the refinement, but I don&#39;t know how to. For NCS, I should define a region that are the same in both monomers? Should I use tight or loose restraints?  For TLS, I don&#39;t have a clue. <br>
<br>      I don&#39;t know whether I am doing the right thing. I did molecular replacement (it seems I got the correct solution), then rigid body refinement, then a couple of restrained refinement right after the rigid body (why the R factors go down even that I haven&#39;t done anything to the structure?), then I walked through the amino acids in coot and did whatever I could. Actually since my DNA and protein helix are longer than the ones in the model, so I built them myself.<br>
<br><br>      Any suggestion is welcome!<br><br><br>Xun<br>      <br><br><br clear="all"><br>-- <br>Department of Molecular and Structural Biochemistry<br>North Carolina State University<br>