Key issues in the computational simulation of GPCR function: representation of loop domains.

Some key concerns raised by molecular modeling and computational simulation of functional mechanisms for membrane proteins are discussed and illustrated for members of the family of G protein coupled receptors (GPCRs). Of particular importance are issues related to the modeling and computational treatment of loop regions. These are demonstrated here with results from different levels of computational simulations applied to the structures of rhodopsin and a model of the 5-HT2A serotonin receptor, 5-HT2AR. First, comparative Molecular Dynamics (MD) simulations are reported for rhodopsin in vacuum and embedded in an explicit representation of the membrane and water environment. It is shown that in spite of a partial accounting of solvent screening effects by neutralization of charged side chains, vacuum MD simulations can lead to severe distortions of the loop structures. The primary source of the distortion appears to be formation of artifactual H-bonds, as has been repeatedly observed in vacuum simulations. To address such shortcomings, a recently proposed approach that has been developed for calculating the structure of segments that connect elements of secondary structure with known coordinates, is applied to 5-HT2AR to obtain an initial representation of the loops connecting the transmembrane (TM) helices. The approach consists of a simulated annealing combined with biased scaled collective variables Monte Carlo technique, and is applied to loops connecting the TM segments on both the extra-cellular and the cytoplasmic sides of the receptor. Although this initial calculation treats the loops as independent structural entities, the final structure exhibits a number of interloop interactions that may have functional significance. Finally, it is shown here that in the case where a given loop from two different GPCRs (here rhodopsin and 5-HT2AR) has approximately the same length and some degree of sequence identity, the fold adopted by the loops can be similar. Thus, in such special cases homology modeling might be used to obtain initial structures of these loops. Notably, however, all other loops in these two receptors appear to be very different in sequence and structure, so that their conformations can be found reliably only by ab initio, energy based methods and not by homology modeling.