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<div dir="auto">Dear Pavel, </div><div dir="auto"><br></div><div dir="auto">Thank you for your prompt response. </div><div dir="auto"><br></div><div dir="auto">We will try and report back on results. </div><div dir="auto"><br></div><div dir="auto">Just a small clarifying question -</div><div dir="auto"><br></div><div dir="auto"><blockquote type="cite" class="gmail_quote" style="padding-left: 0.75ex; border-left-width: 1px; border-left-color: rgb(128, 128, 128); margin: 0px 0px 0px 0.75ex;"><p style="margin-top: 0px; margin-bottom: 0px;">reference_coordinate_restraints.enabled=true</p><p style="margin-top: 0px; margin-bottom: 0px;">reference_coordinate_restraints.selection=water</p><p style="margin-top: 0px; margin-bottom: 0px;">reference_coordinate_restraints.sigma=0.2</p></blockquote></div><div dir="auto"><br></div><div dir="auto"><br></div><div dir="auto">I shall look and set in Real-space refinement protocol? </div><div dir="auto"><br></div><div dir="auto"><br></div><div dir="auto">Many thanks. </div><div dir="auto"><br></div><div dir="auto">Sincerely, </div><div dir="auto">Dmitry</div><div dir="auto"><br></div>
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<p style="color: black; font-size: 10pt; font-family: sans-serif; margin: 8pt 0;">On January 20, 2024 5:25:24 AM Pavel Afonine <pafonine@lbl.gov> wrote:</p>
<blockquote type="cite" class="gmail_quote" style="margin: 0 0 0 0.75ex; border-left: 1px solid #808080; padding-left: 0.75ex;">
Hi Dmitry,<br>
<br>
Here are the considerations regarding water picking in cryo-EM maps:<br>
<br>
Unlike crystallography, where changing the model changes the maps
(e.g., adding more structure improves the maps), in cryo-EM,
changing the model does not change the map. Therefore, tasks such as
refining B factors and adding water are best done using the most
complete and accurate atomic model of the macromolecule, preferably
towards the end.<br>
<br>
<p>Once you have added water, you need to run a round of refinement
and check all water manually. While automated tools generally add
water reasonably well, there is no 100% guarantee that all water
will be correct. Typically, you perform this step once towards the
end.</p>
<p><br>
</p>
<p>"Flying water" seems to be an issue indeed. While I'm working on
the solution, a quick and easy workaround for you to use now is to
restrain water positions to their original coordinates using weak
harmonic restraints. Keywords you'll need:</p>
<p><br>
</p>
<p>reference_coordinate_restraints.enabled=true</p>
<p>reference_coordinate_restraints.selection=water</p>
<p>reference_coordinate_restraints.sigma=0.2</p>
<p><br>
</p>
<p>(also can be set up in the GUI).</p>
<p><br>
</p>
<p>Let me know if you have any questions or need help!<br>
</p>
<p><br>
</p>
<p>Good luck!<br>
</p>
Pavel
<p><br>
</p>
<div class="moz-cite-prefix">On 1/19/24 03:33, Dmitry Semchonok
wrote:<br>
</div>
<blockquote type="cite" cite="mid:AM7PR09MB39899DC7F36600BBDE17CCADF3702@AM7PR09MB3989.eurprd09.prod.outlook.com">
<p class="elementToProof" style="margin-top: 0px; margin-bottom: 0px;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Dear
colleagues, </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p class="elementToProof" style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Please
share your experience with how you handle the water molecules.</span></p>
<p class="elementToProof" style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p class="elementToProof" style="margin: 0cm 0cm 0cm 36pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">So,
we have a refined model and added the water molecules using
the Douse protocol. So, should we refine the model again
afterwards? Or is the model considered ready? </span></p>
<p style="margin: 0cm 0cm 8pt;" class="elementToProof"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p style="margin: 0cm 0cm 8pt;" class="elementToProof"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">We
tried to refine our model with water, but then the water
molecules flew away from the places. </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Please
let me know what is the correct way to proceed. </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Many
thanks for considering my request. </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);"> </span></p>
<p style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Sincerely, </span></p>
<p class="elementToProof" style="margin: 0cm 0cm 8pt;"><span style="font-family: Aptos, sans-serif; font-size: 12pt; color: rgb(0, 0, 0);">Dmitry
</span></p>
<br>
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