Hi. I have a protein structure, GPCR with 400 residues approx. The EC loop contains around 70 residues which makes it so flexible and they are not my attraction of studies, hence i hardly need it.
How is possible to clip the EC loop?
1. How to clip it?
2. Is there any consideration before I clipping?
Please help me with your suggesstions.
I would do it the same way I do it to design an expression construct with a shortened loop - searching for a shorter loop structure that will connect to the base structure without strain, then optimize its sequence using Rosetta.
Hi Dr Honegger
Thank you for your generous advice. with my little knowledge on biological systems, this is almost complicated for me again. Can you precisely help me out with that?
Thank you
Hi Dr Kumar
I was actually studying dimerization, where loop doesn't play any major role. The structure is modeled and refined using dynamics. Unfortunately the loop region is so large and hence flexible to overrun into the membrane region. Anything can be done regarding this?
Which of the three extracellular loop are you talking about?. These two loops are very short in most type A GPCRs, but some type B receptors have long insertions, in some cases representing splicing variants.
The first and third loop you can probably simply replace by a shorter loop from a related GPCR, as long as the flanking helices can be aligned well. Or you can use something like the iMOT server http://caps.ncbs.res.in/imot/iMOTserver.html, giving it the top half of the flanking helices without the connecting loop as search motif, and let it search the structure database for motifs in which two helices with the same relative orientation are connected by a loop.
The second extracellular loop of a GPCR is a bit more complex, as it usually contains two disulfide bridges, one of which connects the loop to the framework of the seven helix bundle, the other lies within the loop, but one Cys is very close to one in the first disulfide bridge. You have to look at your actual structure to see which segments you can cut out to shorten the loop. Alterations of this loop have a high probability of affecting ligand affinity.
If your loop can "overrun into the membrane region", are you simulating the receptor in absence of a membrane or at least lipid micelle? I am highly doubtful that you could get any biologically significant insights into receptor dimerization in such a system! After all, in an aqueous system, the main driving force of interaction is the reduction of solvent exposed hydrophobic surface, which is not a factor in the hydrophobic interior of the membrane.
Dr Annemarie
Thank you
First I apologize for a wrong information I have shared above. Its cytoplasmic loop (3). The protein structure is modelled using I-tasser and the crystal structure is not available though.
The modelled structure itself is having its loop parallel TM domains.
Protein is Vasopressin 1b
Thank you
- Badges
- Science topic