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<p class="MsoNormal">Hi all,<o:p></o:p></p>
<p class="MsoNormal">Have run into an issue with a structure I am refining (3.2A). The temperature factors for the refined solvent molecules show a very weird distribution with half having B’s below 10 and the remainder around 30-50 (I have modelled around
30 solvent in total) – see excerpt below. Average B for protein chains is around 45 and other “solvent” heterogroups (lipid) have B’s around 30-70. The solvent is not part of a TLS group (single TLS group per protein chain). Refining with autoncs / TLSalternate<o:p></o:p></p>
<p class="MsoNormal"><o:p> </o:p></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10043 O HOH C 1 37.384 85.915 48.227 1.00 3.00 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10044 O HOH C 2 41.165 85.554 55.195 1.00 33.13 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10045 O HOH C 3 38.207 79.024 102.082 1.00 40.10 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10046 O HOH C 4 31.928 83.353 106.446 1.00 3.00 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10047 O HOH C 5 59.895 83.126 112.069 1.00 53.13 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10048 O HOH C 6 71.980 110.118 85.303 1.00 41.10 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10049 O HOH C 7 35.067 77.191 94.667 1.00 23.21 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10050 O HOH C 8 58.834 101.563 100.861 1.00 7.26 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10051 O HOH C 9 62.444 104.317 96.908 1.00 3.00 O<o:p></o:p></span></p>
<p class="MsoNormal"><span style="font-family:"Courier New"">HETATM10052 O HOH C 10 31.634 95.839 49.139 1.00 6.41 O<o:p></o:p></span></p>
<p class="MsoNormal">…….<o:p></o:p></p>
<p class="MsoNormal"><o:p> </o:p></p>
<p class="MsoNormal">Solvent peaks return to +fofc maps if I leave them out. If I reset B’s to 30.0 and re-refine, they converge to similar values. I am assuming 3.00 is a minimum Bvalue allowed (similar to Bmax of 300) – oddly it is always the same solvent
molecules that hit 3.00. I do not think there are likely to be many additional ions about – I do have anomalous peaks for calcium sites (whose B’s are in the range 25-40). I am at the end of rebuilding with Rwork/Rfree of 25.0/27.0, fom 0.8.<o:p></o:p></p>
<p class="MsoNormal"><o:p> </o:p></p>
<p class="MsoNormal">Any reason for this behaviour (maybe I am trying to push my luck refining B’s but BUSTER has produced/refined sensible solvent molecules in the past with other datasets at similar resolutions). Hints for what to look for to diagnose the
problem?<o:p></o:p></p>
<p class="MsoNormal"><o:p> </o:p></p>
<p class="MsoNormal">Suggestions welcome,<o:p></o:p></p>
<p class="MsoNormal">Many thanks,<o:p></o:p></p>
<p class="MsoNormal">Ashley<o:p></o:p></p>
<p class="MsoNormal">--<o:p></o:p></p>
<p class="MsoNormal">Dr. Ashley Pike<o:p></o:p></p>
<p class="MsoNormal"><o:p> </o:p></p>
<p class="MsoNormal">Membrane Protein Crystallography | Structural Genomics Consortium | University of Oxford<o:p></o:p></p>
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