Try a smaller integration timestep while you equilibrate. Set nstxout = 1 and look closely at the region that starts to break. Try without constraints. Alternatively, simplify/change the problem by (say) stripping all the crystal waters before adding, or solvating only one copy of the protein (to diagnose topology issues).<br><br>Mark<br><br>----- Original Message -----<br>From: Itamar Kass &lt;itamar.kass@monash.edu&gt;<br>Date: Monday, October 18, 2010 11:14<br>Subject: [gmx-users] Problems when simulating a protein in its crystal        matrix.<br>To: Discussion list for GROMACS users &lt;gmx-users@gromacs.org&gt;<br><br> <span><p>         <meta http-equiv="content-type" content="text/html; charset=ISO-8859-1">   <table><tbody><tr><td text="#000000" bgcolor="#ffffff"><p>     <style>@font-face {   font-family: "Courier New"; }@font-face {   font-family: "Cambria"; }p.MsoNormal, li.MsoNormal, div.MsoNormal { margin: 0cm 0cm 0.0001pt; font-size: 12pt; font-family: "Times New Roman"; }div.Section1 { page: Section1; }</style>     </p><p class="MsoNormal"><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font>Hi All,</p>     <p class="MsoNormal">&nbsp;</p>     <p class="MsoNormal">I wish to simulate a protein in its crystal       form. The       crystal is a triclinic box with a C2 symmetry. <span style="">&nbsp;</span>I       have built the unit cell, which has 4 copies of the       protein, and add some water on top of the ones in the pdb. Then       minimized and run positional restrained MD       followed by a non restrained MD. The system crashed after ~600ps       reporting of LINCS errors.<br>     </p>     <p class="MsoNormal"><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font><br><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font>       Now I       have minimized it before running; I have checked the topology and       could not       find any error; I have also tried to run it in shake and the       system crashed as       well; I also run the system in NVT without any success.</p>     <p class="MsoNormal">&nbsp;</p>     <p class="MsoNormal">I guess this is a symptom to something like       pressure       problems, PBC issues or bad topology, but I could not find it. I       also looked       for an isolated water molecule which might have caused it, without       any success.       So I hope someone else can come with the bright idea I don’t have.</p>     <p class="MsoNormal">&nbsp;</p>     <p class="MsoNormal">The rate of water and protein is ~1 protein to       300 water       molecules, In addition I add NaCl to neutralise the system and       bring it to 0.1M       concentration. </p>     <p class="MsoNormal">&nbsp;</p>     <p class="MsoNormal">Also here is my mdp file:</p>     <p class="MsoNormal"><span style="">integrator<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>md</span></p>     <p class="MsoNormal"><span style="">dt<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>0.002<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span>; ps         !</span></p>     <p class="MsoNormal"><span style="">nsteps<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>2500000</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; OPTIONS FOR BONDS</span></p>     <p class="MsoNormal"><span style="">; Constrain control</span></p>     <p class="MsoNormal"><span style="">constraints<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>all-bonds</span></p>     <p class="MsoNormal"><span style="">; Do not constrain         the start configuration</span></p>     <p class="MsoNormal"><span style="">continuation<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span>= yes</span></p>     <p class="MsoNormal"><span style="">; Type of constraint         algorithm</span></p>     <p class="MsoNormal"><span style="">constraint-algorithm<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span>=<span style="">&nbsp; </span>lincs</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; OUTPUT CONTROL         OPTIONS</span></p>     <p class="MsoNormal"><span style="">; Output frequency         for coords (x), velocities (v) and forces (f)</span></p>     <p class="MsoNormal"><span style="">nstxout<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>100000</span></p>     <p class="MsoNormal"><span style="">nstvout<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>100000</span></p>     <p class="MsoNormal"><span style="">nstfout<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>100000</span></p>     <p class="MsoNormal"><span style="">; Output frequency         and precision for xtc file</span></p>     <p class="MsoNormal"><span style="">nstxtcout<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>2500</span></p>     <p class="MsoNormal"><span style="">xtc-precision<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>1000</span></p>     <p class="MsoNormal"><span style="">; Energy monitoring</span></p>     <p class="MsoNormal"><span style="">energygrps<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>Protein</span></p>     <p class="MsoNormal"><span style="">nstenergy<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>1000</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; NEIGHBORSEARCHING         PARAMETERS</span></p>     <p class="MsoNormal"><span style="">; nblist update         frequency</span></p>     <p class="MsoNormal"><span style="">nstlist<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>5 </span></p>     <p class="MsoNormal"><span style="">; ns algorithm         (simple or grid)</span></p>     <p class="MsoNormal"><span style="">ns-type<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>Grid</span></p>     <p class="MsoNormal"><span style="">; nblist         cut-off<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">rlist<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>0.8</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; OPTIONS FOR ELECTROSTATICS         AND VDW</span></p>     <p class="MsoNormal"><span style="">; Method for doing         electrostatics</span></p>     <p class="MsoNormal"><span style="">coulombtype<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= Reaction-Field<span style="">&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">rcoulomb <span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;</span>= 1.4 </span></p>     <p class="MsoNormal"><span style="">epsilon_rf<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp;         </span> = 62<span style="">&nbsp;&nbsp;&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">vdw-type<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= Cut-off</span></p>     <p class="MsoNormal"><span style="">; cut-off         lengths<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span></span></p>     <p class="MsoNormal"><span style="">rvdw-switch<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= 0</span></p>     <p class="MsoNormal"><span style="">rvdw<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= 1.4</span></p>     <p class="MsoNormal"><span style="">; Apply long range         dispersion corrections for Energy and Pressure</span></p>     <p class="MsoNormal"><span style="">DispCorr<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= no</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; OPTIONS FOR WEAK         COUPLING ALGORITHMS</span></p>     <p class="MsoNormal"><span style="">; Temperature         coupling<span style="">&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">tcoupl<span style="">&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</span>=         berendsen</span></p>     <p class="MsoNormal"><span style="">; Groups to couple         separately, time constant (ps) and reference temperature (K)</span></p>     <p class="MsoNormal"><span style="">tc-grps<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= Protein<span style="">&nbsp;&nbsp; </span>solvent<span style="">&nbsp;         </span></span></p>     <p class="MsoNormal"><span style="">tau-t<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= 0.1<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span>0.1       </span></p>     <p class="MsoNormal"><span style="">ref-t<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= 300<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span>300<span style="">&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">; Pressure coupling<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">Pcoupl<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= no</span></p>     <p class="MsoNormal"><span style="">Pcoupltype<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>= isotropic</span></p>     <p class="MsoNormal"><span style="">; Time constant         (ps), compressibility (1/bar) and reference P (bar)</span></p>     <p class="MsoNormal"><span style="">tau_p<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>1.0<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">compressibility<span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>4.5e-5<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span></span></p>     <p class="MsoNormal"><span style="">ref_p<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span><span style="">&nbsp;&nbsp;&nbsp;&nbsp; </span><span style="">&nbsp;</span>=<span style="">&nbsp;         </span>1.0<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; </span></span></p>     <p class="MsoNormal"><span style="">; Do not generate velocities</span></p>     <p class="MsoNormal"><span style="">gen_vel<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>no</span></p>     <p class="MsoNormal"><span style="">&nbsp;</span></p>     <p class="MsoNormal"><span style="">; Center of mass         control</span></p>     <p class="MsoNormal"><span style="">nstcomm<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>10000</span></p>     <p class="MsoNormal"><span style="">; Periodic boundary         conditions </span></p>     <p class="MsoNormal"><span style="">pbc<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>xyz</span></p>     <p class="MsoNormal"><span style="">; Mode for center of         mass motion removal</span></p>     <p class="MsoNormal"><span style="">comm-mode<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>Linear</span></p>     <p class="MsoNormal"><span style="">; Groups for center         of mass motion removal</span></p>     <p class="MsoNormal"><span style="">comm-grps<span style="">&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;         </span>=<span style="">&nbsp; </span>system</span></p>     <p class="MsoNormal">&nbsp;<br>     </p>     <p class="MsoNormal"><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font>Thanks in advance,<br><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font>       Itamar.<br>     </p>     <pre class="moz-signature" cols="72"><font style="font-style: normal; font-weight: normal; background-color: rgb(245, 248, 240); font-size: 14px;">&gt; </font>--  &gt;  &gt;  &gt; "In theory, there is no difference between theory and practice. But, in practice, there is." - Jan L.A. van de Snepscheut &gt;  &gt; =========================================== &gt; | Itamar Kass, Ph.D. &gt; | Postdoctoral Research Fellow &gt; | &gt; | Department of Biochemistry and Molecular Biology &gt; | Building 77 Clayton Campus &gt; | Wellington Road &gt; | Monash University, &gt; | Victoria 3800 &gt; | Australia &gt; | &gt; | Tel: +61 3 9902 9376 &gt; | Fax: +61 3 9902 9500 &gt; | E-mail: <a href="javascript:main.compose('new', 't=Itamar.Kass@monash.edu')">Itamar.Kass@monash.edu</a> &gt; ============================================</pre>   </td></tr></tbody></table> </p></span> &gt; -- <br>&gt; gmx-users mailing list&nbsp;&nbsp;&nbsp; gmx-users@gromacs.org<br>&gt; http://lists.gromacs.org/mailman/listinfo/gmx-users<br>&gt; Please search the archive at <br>&gt; http://www.gromacs.org/Support/Mailing_Lists/Search before posting!<br>&gt; Please don't post (un)subscribe requests to the list. Use the <br>&gt; www interface or send it to gmx-users-request@gromacs.org.<br>&gt; Can't post? Read http://www.gromacs.org/Support/Mailing_Lists