<HTML><BODY style="word-wrap: break-word; -khtml-nbsp-mode: space; -khtml-line-break: after-white-space; ">Hi all,<DIV><BR class="khtml-block-placeholder"></DIV><DIV>I have a structure refined to 2.5 A. After simulated annealing (start temp = 5000, cool rate = 25, final temp = 100, include TLS refinement and NCS restraint), my R and Rfree are 24.6% and 28.7% respectively. I then turned off the SA in the generated def file and carried on the combined refinement. My R and Rfree went up to 25% and 29.2% respectively. Why did the Rfree go up after refinement?</DIV><DIV><BR class="khtml-block-placeholder"></DIV><DIV>In the remarks of the output pdb file, I have one line, "REMARK twinned : True". What does this mean? </DIV><DIV><BR class="khtml-block-placeholder"></DIV><DIV>The RMSD of bonds and angles after either SA or combined refinement are about 0.006 and 0.851 respectively. Don't you guys think the geometry restraints are too tight? How do I change the weight in PHENIX?</DIV><DIV><BR class="khtml-block-placeholder"></DIV><DIV>Thanks.</DIV><DIV><BR class="khtml-block-placeholder"></DIV><DIV>Jianghai</DIV><DIV><BR><DIV> <SPAN class="Apple-style-span" style="border-collapse: separate; border-spacing: 0px 0px; color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; text-align: auto; -khtml-text-decorations-in-effect: none; text-indent: 0px; -apple-text-size-adjust: auto; text-transform: none; orphans: 2; white-space: normal; widows: 2; word-spacing: 0px; "><SPAN class="Apple-style-span" style="border-collapse: separate; border-spacing: 0px 0px; color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; text-align: auto; -khtml-text-decorations-in-effect: none; text-indent: 0px; -apple-text-size-adjust: auto; text-transform: none; orphans: 2; white-space: normal; widows: 2; word-spacing: 0px; "><SPAN class="Apple-style-span" style="border-collapse: separate; border-spacing: 0px 0px; color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; text-align: auto; -khtml-text-decorations-in-effect: none; text-indent: 0px; -apple-text-size-adjust: auto; text-transform: none; orphans: 2; white-space: normal; widows: 2; word-spacing: 0px; "><SPAN class="Apple-style-span" style="border-collapse: separate; border-spacing: 0px 0px; color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; text-align: auto; -khtml-text-decorations-in-effect: none; text-indent: 0px; -apple-text-size-adjust: auto; text-transform: none; orphans: 2; white-space: normal; widows: 2; word-spacing: 0px; "><SPAN class="Apple-style-span" style="border-collapse: separate; border-spacing: 0px 0px; color: rgb(0, 0, 0); font-family: Helvetica; font-size: 12px; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; text-align: auto; -khtml-text-decorations-in-effect: none; text-indent: 0px; -apple-text-size-adjust: auto; text-transform: none; orphans: 2; white-space: normal; widows: 2; word-spacing: 0px; "><DIV>+++++++++++++++++++++++++++++++</DIV><DIV>Jianghai Zhu, Ph.D</DIV><DIV>CBR Institute for Biomedical Research</DIV><DIV>Department of Pathology</DIV><DIV>Harvard Medical School</DIV><DIV>200 Longwood Ave., Boston, MA 02115</DIV><DIV>Ph: 617-278-3211</DIV><DIV>Fx: 618-278-3030</DIV><DIV>+++++++++++++++++++++++++++++++</DIV><DIV><BR class="khtml-block-placeholder"></DIV><BR class="Apple-interchange-newline"></SPAN></SPAN></SPAN></SPAN></SPAN> </DIV><BR></DIV></BODY></HTML>