Hi all
I am new to modeller and i'm currently trying to build a model of a GPCR using the bovine rhodopsin structure as template. I have notice that in the middle of one of a region corresponding to a transmembrane segment, there is a very unambiguous gap of two residue in the sequence alignment.
Two model of this receptor have already been published and in both cases, authors have used sequence alignment without gaps, which means that half the residues of this TM are clearly not correctly aligned with the template.
I am confuse of how I should handle this gap to build my model. Using a 2 residues helix-gapped alignment in modeller would result in a model with - ugly coil in the middle of the helix - a switch from one helix face to the opposite of amino acids - a longer loop next to the TM (or maybe a longer helix ?) On the contrary, ignoring the gap and using a wrong alignment result in a much better looking helix.
Should I deliberately ignore the sequence similarity to provide modeller an alignment with no gap in the TM ?
Best regards, Guillaume LETELLIER
Hi,
May be you would like to have a look to MODELLER's ALPHA restraints. They take the helical gaps into consideration very nicely. Prevents kinking and keeps the faces nicely.
Hope it helps.
br,
Vivek Sharma
On 2/2/06, Guillaume LETELLIER guillaume.letellier@cea.fr wrote: > > Hi all > > I am new to modeller and i'm currently trying to build a model of a GPCR > using the bovine rhodopsin structure as template. > I have notice that in the middle of one of a region corresponding to a > transmembrane segment, there is a very unambiguous gap of two residue in > the sequence alignment. > > Two model of this receptor have already been published and in both > cases, authors have used sequence alignment without gaps, which means > that half the residues of this TM are clearly not correctly aligned with > the template. > > I am confuse of how I should handle this gap to build my model. > Using a 2 residues helix-gapped alignment in modeller would result in a > model with > - ugly coil in the middle of the helix > - a switch from one helix face to the opposite of amino acids > - a longer loop next to the TM (or maybe a longer helix ?) > On the contrary, ignoring the gap and using a wrong alignment result in > a much better looking helix. > > Should I deliberately ignore the sequence similarity to provide modeller > an alignment with no gap in the TM ? > > > Best regards, > Guillaume LETELLIER > > > _______________________________________________ > modeller_usage mailing list > modeller_usage@salilab.org > http://salilab.org/mailman/listinfo/modeller_usage > > > >
Vivek Sharma wrote: > Hi, > > May be you would like to have a look to MODELLER's ALPHA restraints. > They take the helical gaps into consideration very nicely. Prevents > kinking and keeps the faces nicely. > > Hope it helps. Indeed it helped ! I had already try the alpha constraint, but i was applying it only to the gapped residues. I just realized that modeller works much better when applying the constraint to the whole helix length.
Thank you very much.
> Hi Guillaume, > I'm a little confuse about your explanation...the gap is real or not? > Hi Helena, and thank you for answering my unclear question. > Probably you will need to write to those who modelled the structures > before to ask about this gap. i think is really unusual that someone > could published structures that have wrong alignments... This is right, i should probably have started here. I was very surprised by these data, and i've checked every possible confusion, but there's no doubt i'm working on the same structure and sequences. For me, the gap in the helix is really obvious, although it seems to have been ignored in previous papers.
here's the 'reference' alignment of the helix sequence found in the literatture : > PLNYILLNLAVADLFMVFGGFTTTLYTSLH > VNNYFLLSLACADLIIGTFSMNLYTTYLLM ** ** ** *** . *
and here's the one I use : > PLNYILLNLAVADLFMVFGGFTTTLYTSLH-- > VNNYFLLSLACADL--IIGTFSMNLYTTYLLM ** ** ** *** : * *: ***:
...
Guillaume
Hi,
Just to get this out of the way, I am not an expert on GPCR and have never attempted to model them. Still, I suggest you look at my arguments below.
At 09:03 AM 2/2/2006, you wrote: >>Hi Guillaume, >>I'm a little confuse about your explanation...the gap is real or not? >> >Hi Helena, and thank you for answering my unclear question. >>Probably you will need to write to those who modelled the structures >>before to ask about this gap. i think is really unusual that someone >>could published structures that have wrong alignments... >This is right, i should probably have started here. >I was very surprised by these data, and i've checked every possible >confusion, but there's no doubt i'm working on the same structure and >sequences. >For me, the gap in the helix is really obvious, although it seems to have >been ignored in previous papers. > >here's the 'reference' alignment of the helix sequence found in the >literatture : > > PLNYILLNLAVADLFMVFGGFTTTLYTSLH > > VNNYFLLSLACADLIIGTFSMNLYTTYLLM > ** ** ** *** . * > >and here's the one I use : > > PLNYILLNLAVADLFMVFGGFTTTLYTSLH-- > > VNNYFLLSLACADL--IIGTFSMNLYTTYLLM > ** ** ** *** : * *: ***:
As a general argument, just because an alignment has more true matches does not mean that it is correct. There are penalties associated with those two gaps you introduced that offset the gain from exact residue matching. Not knowing exactly which sequence you are modeling, I submitted P11229 (ACM1_HUMAN) to the SUPERFAMILY server and the alignment come back with no gaps in that region. Take a look at:
http://www.supfam.org/SUPERFAMILY/cgi-bin/gene.cgi?genome=up;seqid=P11229
and follow the "Align" link. Obviously, the same can be done for any protein for which you know the SwissProt number by simply inserting it at the end of the link above. Not to get into prolonged argument here, I'll just say that I would trust SUPERFAMILY-based alignment over any human-derived alignment unless there is a very good reason to believe that the server alignment is wrong. And breaking a long helix to make few extra positive matches usually doesn't qualify as a good reason, especially because it forces you to make a compensatory two-residue gap later in the model. But don't take my word for it, try modeling the protein with and without those gaps and see which model evaluates better.
Hope this helps,
Mensur
========================================================================== | Mensur Dlakic, PhD | Tel: (406) 994-6576 | | Department of Microbiology | Fax: (406) 994-4926 | | Montana State University | | | 109 Lewis Hall, P.O. Box 173520 | http://myprofile.cos.com/mensur | | Bozeman, MT 59717-3520 | E-mail: mdlakic@montana.edu | ==========================================================================
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Guillaume LETELLIER -
Mensur Dlakic -
Vivek Sharma