We have a problem building a 2-domain protein based on multiple (8) other proteins. We put a slash between the two domains so that Modeller knows not to connect the termini. We have no problems with large distances between equivalent CA's of different proteins. the only unusual aspect we notice is that the pair of warnings (?) "drmsq1__W> n<2 and drmsq3__W> n<2" occurs 19 times. No error message occurs, but no model is produced either. Please find attached part of the output. Hope this is enough for the experts on ths list to give us hints as to how to resolve this issue.
Thanks a million,
Pieter Stouten pieter.stouten@dupontmerck.com DuPont Merck - Computer Aided Drug Design Group Web: http://www.halcyon.com/stouten/
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chkaln___> Checking pairwise structural superpositions: Equivalent CA pairs with distance difference larger than 6.0 angstroms:
ALN_POS TMPL1 TMPL2 RID1 RID2 NAM1 NAM2 DIST ---------------------------------------------------- drmsq1__W> n<2 drmsq3__W> n<2
[... repeated 19 times ... /PFWS]
chkaln___> Checking structure-sequence alignments: Implied target CA(i)-CA(i+1) distances longer than 8.0 angstroms:
ALN_POS TMPL RID1 RID2 NAM1 NAM2 DIST ----------------------------------------------
<< end of CHECK_ALIGNMENT.
TOP______> 109 94 CALL ROUTINE = BUILD_METHOD
TOP______> 110 66 READ_ALIGNMENT FILE = ALNFILE, ALIGN_CODES = KNOWNS
[...]
TOP______> 111 67 MALIGN3D FIT = OFF, GAP_PENALTIES_3D = 0 4
malign3d_> Initial framework positions: 9 fit2xyz_E> number of equivalent positions < 3: 0 recover__> MODELLER_STATUS >= CONTROL: 1 1
Dear Pieter!
Initial alignment is porbably bad which is indicated in diagnostics of CHECK_ALIGNMENT and MALIGN3D. Please check your alignment manually or use FIT = on in MALIGN3D to produce correct alignment. If it won't help, then please send me .top, .pdb, alignment files and other necessary input files in separate attachments or as a tar.gz archive.
Happy landings,
Azat
Pieter Stouten wrote: > > We have a problem building a 2-domain protein based on multiple (8) other > proteins. We put a slash between the two domains so that Modeller knows not > to connect the termini. We have no problems with large distances between > equivalent CA's of different proteins. the only unusual aspect we notice is > that the pair of warnings (?) "drmsq1__W> n<2 and drmsq3__W> n<2" occurs 19 > times. No error message occurs, but no model is produced either. Please > find attached part of the output. Hope this is enough for the experts on > ths list to give us hints as to how to resolve this issue. > > Thanks a million, > > Pieter Stouten pieter.stouten@dupontmerck.com > DuPont Merck - Computer Aided Drug Design Group > Web: http://www.halcyon.com/stouten/ > > ---------------------------------------------------------------------------- > > chkaln___> Checking pairwise structural superpositions: > Equivalent CA pairs with distance difference larger than 6.0 > angstroms: > > ALN_POS TMPL1 TMPL2 RID1 RID2 NAM1 NAM2 DIST > ---------------------------------------------------- > drmsq1__W> n<2 > drmsq3__W> n<2 > > [... repeated 19 times ... /PFWS] > > chkaln___> Checking structure-sequence alignments: > Implied target CA(i)-CA(i+1) distances longer than 8.0 angstroms: > > ALN_POS TMPL RID1 RID2 NAM1 NAM2 DIST > ---------------------------------------------- > > << end of CHECK_ALIGNMENT. > > TOP______> 109 94 CALL ROUTINE = BUILD_METHOD > > TOP______> 110 66 READ_ALIGNMENT FILE = ALNFILE, ALIGN_CODES = KNOWNS > > [...] > > TOP______> 111 67 MALIGN3D FIT = OFF, GAP_PENALTIES_3D = 0 4 > > malign3d_> Initial framework positions: 9 > fit2xyz_E> number of equivalent positions < 3: 0 > recover__> MODELLER_STATUS >= CONTROL: 1 1
-- - Dr. Azat Badretdinov - The Rockefeller Univ, Box 270 - 1230 York Ave, New York NY 10021, USA - Phone: (212) 327 7206 - Fax: (212) 327 7540 - E-mail: azat@salilab.org - WWW/URL: http://salilab.org/~azat
Hello Azat,
Thanks for your mail of 98/06/23 at 8:43 -0400.
>Initial alignment is probably bad which is indicated in diagnostics of >CHECK_ALIGNMENT and MALIGN3D. > Please bear with me for a moment: although I have used "Modeller" before, I never ran into problems so I never paid much attention to the diagnostics messages. However, CHECK_ALIGNMENT indicated no (zero) equivalent CA pairs with distance difference larger than 6.0 angstroms and no (zero) implied target CA(i)-CA(i+1) distances longer than 8.0 angstroms. It did list 19 occurrences of the statements "drmsq1__W> n<2" and "drmsq3__W> n<2". We had seen these messages before (never 19 times, though), but still obtained good models. My deduction is that the alignment should be good enough to produce a model, so I cannot explain why it does not do so. Also, modelling the individual domains with the exact same sequence alignments did not lead to any problems.
>Please check your alignment manually > Done twice already.
>or use FIT = on in MALIGN3D to produce correct alignment. > We might give that a shot, just to see if that solves the problem (not because we think a better alignment would be produced <g>).
>If it won't help, then please send me .top, .pdb, alignment files and >other necessary input files > I am afraid I cannot do that as the files contain proprietary information. I have sent the lot off to MSI, though. I hope they can help. Thanks for the offer, anyway.
Cheers,
Pieter
Pieter Stouten || Nothing shocks me; Computer Aided Drug Design Group || The DuPont Merck Pharmaceutical Company || I am a scientist! P.O. Box 80500, Wilmington, DE 19880-0500 || Phone: +1 (302) 695 3515 || -- Fax: +1 (302) 695 9090 || Internet: pieter.stouten@dupontmerck.com || Indiana Jones Web: http://www.halcyon.com/stouten/ ||
Dear Pieter;
If I read correctly the last 3 sentences of the log file you obtained:
> malign3d_> Initial framework positions: 9 > fit2xyz_E> number of equivalent positions < 3: 0 > recover__> MODELLER_STATUS >= CONTROL: 1 1
It seems that you tryed to used malign3d with 9 sequences but you told us that you have only 8 structures. That is probably the problem, because Malign3D works only with structures. Here you included the sequence to model, that crashed Modeller. Please Check you top file again.
Eric
-- Eric FEYFANT Laboratory of Molecular Biophysics (SALI Lab) The Rockefeller Univ, Box 270 1230 York Ave | New York NY 10021 | USA | Phone: (212) 327 7206 | Fax: (212) 327 7540 E-mail: eric@flute.rockefeller.edu www-homepage:http://salilab.org/~eric/
Hello Eric,
>If I read correctly the last 3 sentences of the log file you obtained: > >>malign3d_> Initial framework positions: 9 >>fit2xyz_E> number of equivalent positions < 3: 0 >>recover__> MODELLER_STATUS >= CONTROL: 1 1 > > It seems that you tryed to used malign3d with 9 sequences but you told > us that you have only 8 structures. That is probably the problem, > because Malign3D works only with structures. Here you included the > sequence to model, that crashed Modeller. Please Check you top file > again. > I did. The TOP file only lists 8 known structures. Since no numbers are specified in the TOP file, I do not know how "Modeller" could think there are 9. the output does say that 9 sequences are aligned, which is correct, as the model sequence is aligned with the other 8. I appreciate your help, but unfortunately it does not seem to solve the problem.
BTW, could you please not use HTML in mail? Although my mail reader can handle it, many people can only read plain ASCII. Styled text in e-mail is like a fish on a bicycle.
Thanks again,
Pieter
Pieter Stouten || Computer Aided Drug Design Group || Free advice is The DuPont Merck Pharmaceutical Company || seldom cheap P.O. Box 80500, Wilmington, DE 19880-0500 || Phone: +1 (302) 695 3515 || (Ferengi Rule Of Fax: +1 (302) 695 9090 || Acquisition #59) Internet: pieter.stouten@dupontmerck.com || Web: http://www.halcyon.com/stouten/ ||
I am glad to announce that Tina Yeh at MSI found the culprit. It seems that Modeller aligns all structures to the first one. In our case the first structure (which was not our primary template) had only 9 residues and, therefore, several of the other structures could not be superimposed on it. Making the primary template the first parameter of the "SET KNOWNS" command turned out to be the solution. Nothing else needed to be changed.
Thanks everybody for trying to help us out. It is much appreciated.
Cheers,
Pieter
Pieter Stouten || Computer Aided Drug Design Group || No good deed ever The DuPont Merck Pharmaceutical Company || goes unpunished P.O. Box 80500, Wilmington, DE 19880-0500 || Phone: +1 (302) 695 3515 || (Ferengi Rule Of Fax: +1 (302) 695 9090 || Acquisition #285) Internet: pieter.stouten@dupontmerck.com || Web: http://www.halcyon.com/stouten/ ||
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Azat Badretdinov -
Eric Feyfant -
Pieter Stouten