<html><head><meta http-equiv="Content-Type" content="text/html charset=iso-8859-1"></head><body style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; ">Thanks, Pavel.<div><br></div><div>So you don't think that the difference between Wilson B (66.45 according to phenix) and average B (52 after refinement) is still too large? Where did you take the statistics from? Is that published anywhere?</div><div><br></div><div>Best regards</div><div><br></div><div>Florian</div><div><br><div><div>Am 22.03.2013 um 07:09 schrieb Pavel Afonine &lt;<a href="mailto:pafonine@lbl.gov">pafonine@lbl.gov</a>&gt;:</div><br class="Apple-interchange-newline"><blockquote type="cite">
  
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    <div class="moz-cite-prefix">Hi Florian,<br>
      <br>
      I think it is pretty convincing that refinement went all right and
      the mean B you get is what it actually is. Also I just looked at
      the files that you sent me the other day and I did not notice
      anything suspicious.<br>
      <br>
      Pavel<br>
      <br>
      P.S.: Sorry if you received multiple copies of this email: I was
      trying to attach a tiny image with some PDB statistics but our
      server did not let me do it.<br>
      <br>
      On 3/21/13 1:00 AM, Florian Br�ckner wrote:<br>
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      Dear Pavel
      <div><br>
      </div>
      <div>yes, the refinement did converge. I did 6 macrocycles and in
        separate runs started from different average atomic B-factors of
        32, 66 and 97 keeping the distribution as in the original high
        resolution molecular replacement model from the same crystal
        form (i.e. adding 0, 34 and 65 to all atomic B-factors in the
        starting pdb). In all cases after 6 macrocycles I get an almost
        identical distribution of B-factors and also average B-factor of
        about 52. Also setting all B-factors to the same value of 66
        gave a similar distribution after refinement with an average of
        52. However, in the latter case the R-free was about 2% higher
        compared to starting from the original distribution, but it
        dropped by about 1% during refinement.</div>
      <div><br>
      </div>
      <div>Best regards</div>
      <div><br>
      </div>
      <div>Florian</div>
      <div><br>
      </div>
      <div><br>
        <div>
          <div>Am 21.03.2013 um 06:40 schrieb Pavel Afonine &lt;<a moz-do-not-send="true" href="mailto:pafonine@lbl.gov">pafonine@lbl.gov</a>&gt;:</div>
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              <div class="moz-cite-prefix">Hi Florian,<br>
                <br>
                I'm glad it works now. Regarding difference between
                Wilson B and mean refined B: well, it's hard to quantify
                what's "too low" or "too different". Average refined B
                may deviate from Wilson B and the difference in your
                case does not seem to be terribly large. <br>
                Did the refinement converge? What if you do say 10 or 20
                macro-cycles instead of default 3? Will B-factors
                further change between macro-cycles?<br>
                <br>
                Pavel<br>
                <br>
                On 3/19/13 7:37 AM, Florian Br�ckner wrote:<br>
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                Dear Keitaro and Pavel,
                <div><br>
                </div>
                <div>thank you for your suggestions. I have now used the
                  latest nightly build (1326). With this version, the
                  average B-factor converges at about 52 A**2 and also
                  the distribution is very similar, regardless with what
                  B-factors I start the refinement (I tried average
                  B-factors between 32 and 97). So clearly the behavior
                  of this newest version is different from the version I
                  have been using before (1.8.1 stable release 1168) and
                  also from the latest stable release (1.8.2 stable
                  release 1309), which behaves the same as 1168. To me
                  the B-factors still look a bit too low, since the
                  Wilson B-factor calculated by phenix is 66.45, the
                  Wilson B-factor I determined manually from the linear
                  part of the Wilson plot is 97.</div>
                <div><br>
                </div>
                <div>Best regards</div>
                <div><br>
                </div>
                <div>Florian</div>
                <div><br>
                </div>
                <div><br>
                  <div>
                    <div>Am 19.03.2013 um 00:06 schrieb Keitaro
                      Yamashita &lt;<a moz-do-not-send="true" href="mailto:yamashita@castor.sci.hokudai.ac.jp">yamashita@castor.sci.hokudai.ac.jp</a>&gt;:</div>
                    <br class="Apple-interchange-newline">
                    <blockquote type="cite">Dear Florian,<br>
                      <br>
                      Maybe you are using phenix-dev-1311 or earlier?<br>
                      I am afraid that you have experienced the bug that
                      overall B-factor is<br>
                      not applied to individual atomic B-factors in
                      certain cases, which I<br>
                      have also experienced recently. That bug was fixed
                      in dev-1312 if I<br>
                      recall correctly.<br>
                      <br>
                      Hope it helps,<br>
                      Keitaro<br>
                      <br>
                      2013/3/19 Florian Br�ckner &lt;<a moz-do-not-send="true" href="mailto:florian.brueckner@psi.ch">florian.brueckner@psi.ch</a>&gt;:<br>
                      <blockquote type="cite">Dear all<br>
                        <br>
                        I am refining a structure at 3.2 A with
                        phenix.refine using automatic<br>
                        optimization of target weights, solved by
                        molecular replacement with a 2.1A<br>
                        structure of the same crystal form. Rwork=26%,
                        Rfree=28%. I use individual<br>
                        atomic B-factor refinement. The obtained
                        distribution of B-factors after<br>
                        refinement looks reasonable and quite similar to
                        the distribution in the<br>
                        higher resolution model, which I suppose would
                        be expected, since it is the<br>
                        same crystal form. However, the average B-factor
                        does not change<br>
                        substantially, i.e. if I start with the
                        B-factors from the high resolution<br>
                        model (average B-factor around 32 A**2) after 6
                        macrocycles the average<br>
                        B-factor is still around 32 A**2. If I set all
                        B-factors to the Wilson<br>
                        B-factor determined by phenix.refine (66 A**2),
                        after 6 macrocycles the<br>
                        distribution of B-factors looks again very
                        similar to the high resolution MR<br>
                        model, but the average B-factor is still around
                        65 A**2. What should I make<br>
                        of this? Is this an expected behavior at this
                        resolution, i.e. the fact that<br>
                        the absolute values of B-factors cannot be
                        determined during refinement,<br>
                        although a reasonable distribution can be
                        obtained as judged by plotting<br>
                        average B-factors vs. residue#? Since the
                        average B-factor does not change<br>
                        much during refinement, which average B-factor
                        should I start refinement<br>
                        with to end up with reasonable absolute values?
                        I have manually determined<br>
                        the Wilson B-factor from the linear part of the
                        Wilson plot (4.5-3.2 A) to<br>
                        be 97 A**2. This seems high to me. Would that be
                        expected for the given<br>
                        resolution, given that the 2.1A structure of the
                        same crystal form has 32<br>
                        A**2? I should add, that there is not a
                        substantial effect of the average<br>
                        refined B-factor on Rfree.<br>
                        <br>
                        Thanks for any comments or suggestions.<br>
                        <br>
                        Best regards<br>
                        <br>
                        Florian<br>
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                                            Florian Br�ckner<br>
                                            Laboratory of Biomolecular
                                            Research (LBR)</div>
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                                            Paul Scherrer Institut<br>
                                            CH-5232 Villigen PSI<br>
                                            Switzerland<br>
                                            <br>
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                                            Email:<span class="Apple-converted-space">&nbsp;</span><span class="Apple-tab-span" style="white-space: pre; ">
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reak-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; "><span class="Apple-style-span" style="border-collapse: separate; color: rgb(0, 0, 0); font-family: Helvetica; font-style: normal; font-variant: normal; font-weight: normal; letter-spacing: normal; line-height: normal; orphans: 2; text-indent: 0px; text-transform: none; white-space: normal; widows: 2; word-spacing: 0px; -webkit-border-horizontal-spacing: 0px; -webkit-border-vertical-spacing: 0px; -webkit-text-decorations-in-effect: none; -webkit-text-size-adjust: auto; -webkit-text-stroke-width: 0px; font-size: medium; "><div style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; ">-------------------------------------------------------------</div><div style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; "><br></div><div style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; ">Dr. Florian Br�ckner<br>Laboratory of Biomolecular Research (LBR)</div><div style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; ">OFLG/102<br>Paul Scherrer Institut<br>CH-5232 Villigen PSI<br>Switzerland<br><br></div><div style="word-wrap: break-word; -webkit-nbsp-mode: space; -webkit-line-break: after-white-space; ">Tel.:<span class="Apple-tab-span" style="white-space: pre; ">                </span>+41-(0)56-310-2332<br>Email:<span class="Apple-converted-space">&nbsp;</span><span class="Apple-tab-span" style="white-space: pre; ">        </span><a href="mailto:florian.brueckner@psi.ch">florian.brueckner@psi.ch</a><br><br><br><br></div></span></div></span></div></span></div></span></div></span></div></span></div></span></div></span></span>
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