@article { ISI:000310482800011, title = {Identification of cholesterol binding sites in the serotonin(1A) receptor}, journal = {Journal of Physical Chemistry B}, volume = {116}, number = {43}, year = {2012}, month = {NOV}, pages = {12991-12996}, publisher = {AMER CHEMICAL SOC}, address = {1155 16TH ST, NW, WASHINGTON, DC 20036 USA}, abstract = {

The serotonin(1A) receptor is a representative member of the G protein-coupled receptor (GPCR) superfamily and serves as an important drug target in the development of therapeutic agents for neuropsychiatric disorders. Previous work has shown the requirement of membrane cholesterol in the organization, dynamics, and function of the serotonin(1A) receptor. We show here that membrane cholesterol binds preferentially to certain sites on the serotonin(1A) receptor by performing multiple, long time scale MARTINI coarse-grain molecular dynamics simulations. Interestingly, our results identify the highly conserved cholesterol recognition/interaction amino acid consensus (CRAC) motif on transmembrane helix V as one of the sites with high cholesterol occupancy, thereby confirming its role as a putative cholesterol binding motif. These results represent the first direct evidence for membrane cholesterol binding to specific sites on the serotonin(1A) receptor and represent an important step in our overall understanding of GPCR function in health and disease.

}, issn = {1520-6106}, doi = {10.1021/jp309888u}, author = {Sengupta, Durba and Chattopadhyay, Amitabha} } @article { ISI:000333226900011, title = {Cholesterol modulates the dimer interface of the beta(2)-adrenergic receptor via cholesterol occupancy sites}, journal = {Biophysical Journal}, volume = {106}, number = {6}, year = {2014}, month = {MAR}, pages = {1290-1300}, publisher = {CELL PRESS}, type = {Article}, address = {600 TECHNOLOGY SQUARE, 5TH FLOOR, CAMBRIDGE, MA 02139 USA}, abstract = {

The beta(2)-adrenergic receptor is an important member of the G-protein-coupled receptor (GPCR) superfamily, whose stability and function are modulated by membrane cholesterol. The recent high-resolution crystal structure of the beta(2)-adrenergic receptor revealed the presence of possible cholesterol-binding sites in the receptor. However, the functional relevance of cholesterol binding to the receptor remains unexplored. We used MARTINI coarse-grained molecular-dynamics simulations to explore dimerization of the beta(2)-adrenergic receptor in lipid bilayers containing cholesterol. A novel (to our knowledge) aspect of our results is that receptor dimerization is modulated by membrane cholesterol. We show that cholesterol binds to transmembrane helix IV, and cholesterol occupancy at this site restricts its involvement at the dimer interface. With increasing cholesterol concentration, an increased presence of transmembrane helices I and II, but a reduced presence of transmembrane helix IV, is observed at the dimer interface. To our knowledge, this study is one of the first to explore the correlation between cholesterol occupancy and GPCR organization. Our results indicate that dimer plasticity is relevant not just as an organizational principle but also as a subtle regulatory principle for GPCR function. We believe these results constitute an important step toward designing better drugs for GPCR dimer targets.

}, issn = {0006-3495}, doi = {10.1016/j.bpj.2014.02.002}, author = {Prasanna, Xavier and Chattopadhyay, Amitabha and Sengupta, Durba} } @article { ISI:000369744300004, title = {Differential dynamics of the serotonin(1A) receptor in membrane bilayers of varying cholesterol content revealed by all atom molecular dynamics simulation}, journal = {Molecular Membrane Biology}, volume = {32}, number = {4}, year = {2015}, month = {MAY}, pages = {127-137}, publisher = {TAYLOR \& FRANCIS LTD}, address = {4 PARK SQUARE, MILTON PARK, ABINGDON OX14 4RN, OXON, ENGLAND}, abstract = {

The serotonin(1A) receptor belongs to the superfamily of G protein-coupled receptors (GPCRs) and is a potential drug target in neuropsychiatric disorders. The receptor has been shown to require membrane cholesterol for its organization, dynamics and function. Although recent work suggests a close interaction of cholesterol with the receptor, the structural integrity of the serotonin(1A) receptor in the presence of cholesterol has not been explored. In this work, we have carried out all atom molecular dynamics simulations, totaling to 3s, to analyze the effect of cholesterol on the structure and dynamics of the serotonin(1A) receptor. Our results show that the presence of physiologically relevant concentration of membrane cholesterol alters conformational dynamics of the serotonin(1A) receptor and, on an average lowers conformational fluctuations. Our results show that, in general, transmembrane helix VII is most affected by the absence of membrane cholesterol. These results are in overall agreement with experimental data showing enhancement of GPCR stability in the presence of membrane cholesterol. Our results constitute a molecular level understanding of GPCR-cholesterol interaction, and represent an important step in our overall understanding of GPCR function in health and disease.

}, keywords = {GPCR, Membrane cholesterol, molecular dynamics simulation, serotonin(1A) receptor}, issn = {0968-7688}, doi = {10.3109/09687688.2015.1096971}, author = {Patra, Swarna M. and Chakraborty, Sudip and Shahane, Ganesh and Prasanna, Xavier and Sengupta, Durba and Maiti, Prabal K. and Chattopadhyay, Amitabha} } @article { ISI:000358627400006, title = {Molecular dynamics simulations of GPCR-cholesterol interaction: an emerging paradigm}, journal = {Biochimica Et Biophysica Acta-Biomembranes}, volume = {1848}, number = {9, SI}, year = {2015}, month = {SEP}, pages = {1775-1782}, publisher = {ELSEVIER SCIENCE BV}, address = {PO BOX 211, 1000 AE AMSTERDAM, NETHERLANDS}, abstract = {

G protein-coupled receptors (GPCRs) are the largest class of molecules involved in signal transduction across cell membranes and represent major targets in the development of novel drug candidates. Membrane cholesterol plays an important role in GPCR structure and function. Molecular dynamics simulations have been successful in exploring the effect of cholesterol on the receptor and a general consensus molecular view is emerging. We review here recent molecular dynamics studies at multiple resolutions highlighting the main features of cholesterol-GPCR interaction. Several cholesterol interaction sites have been identified on the receptor that are reminiscent of nonannular sites. These cholesterol hot-spots are highly dynamic and have a microsecond time scale of exchange with the bulk lipids. A few consensus sites (such as the CRAC site) have been identified that correspond to higher cholesterol interaction. Interestingly, high plasticity is observed in the modes of cholesterol interaction and several sites have been suggested to have high cholesterol occupancy. We therefore believe that these cholesterol hot-spots are indicative of {\textquoteleft}high occupancy sites{\textquoteright} rather than {\textquoteleft}binding sites{\textquoteright}. The results suggest that the energy landscape of cholesterol association with GPCRs corresponds to a series of shallow minima interconnected by low barriers. These specific interactions, along with general membrane effects, have been observed to modulate GPCR organization. Membrane cholesterol effects on receptor structure and organization, that in turn influences receptor cross-talk and drug efficacy, represent a new frontier in GPCR research. This article is part of a Special Issue entitled: Lipid-protein interactions. Guest Editors: Amitabha Chattopadhyay and jean-Marie Ruysschaert. (C) 2015 Elsevier B.V. All rights reserved.

}, keywords = {Coarse-grain simulation, CRAC, GPCR, Lipid-receptor interaction, Membrane cholesterol, Receptor dimerization}, issn = {0005-2736}, doi = {10.1016/j.bbamem.2015.03.018}, author = {Sengupta, Durba and Chattopadhyay, Amitabha} } @inbook { ISI:000350416600017, title = {Role of lipid-mediated effects in beta(2)-adrenergic receptor dimerization}, booktitle = {biochemical roles of eukaryotic cell surface macromolecules}, series = {Advances in Experimental Medicine and Biology}, volume = {842}, year = {2015}, pages = {247-261}, publisher = {Springer-Verlag Berlin}, organization = {Springer-Verlag Berlin}, address = {Heidelberger Platz 3, D-14197 Berlin, Germany :}, isbn = {978-3-319-11280-0; 978-3-319-11279-4}, issn = {0065-2598}, doi = {10.1007/978-3-319-11280-0_16}, author = {Prasanna, Xavier and Chattopadhyay, Amitabha and Sengupta, Durba}, editor = {Chakrabarti, A. and Surolia, A.} } @article { ISI:000381562500001, title = {Cholesterol-dependent conformational plasticity in GPCR dimers}, journal = {Scientific Reports}, volume = {6}, year = {2016}, month = {AUG}, pages = {31858}, publisher = {NATURE PUBLISHING GROUP}, address = {MACMILLAN BUILDING, 4 CRINAN ST, LONDON N1 9XW, ENGLAND}, abstract = {

The organization and function of the serotonin1A receptor, an important member of the GPCR family, have been shown to be cholesterol-dependent, although the molecular mechanism is not clear. We performed a comprehensive structural and dynamic analysis of dimerization of the serotonin1A receptor by coarse-grain molecular dynamics simulations totaling 3.6 ms to explore the molecular details of its cholesterol-dependent association. A major finding is that the plasticity and flexibility of the receptor dimers increase with increased cholesterol concentration. In particular, a dimer interface formed by transmembrane helices I-I was found to be sensitive to cholesterol. The modulation of dimer interface appears to arise from a combination of direct cholesterol occupancy and indirect membrane effects. Interestingly, the presence of cholesterol at the dimer interface is correlated with increased dimer plasticity and flexibility. These results represent an important step in characterizing the molecular interactions in GPCR organization with potential relevance to therapeutic interventions.

}, issn = {2045-2322}, doi = {10.1038/srep31858}, author = {Prasanna, Xavier and Sengupta, Durba and Chattopadhyay, Amitabha} } @article { ISI:000375142200228, title = {Conformational dynamics of GPCR dimers is dependent on membrane cholesterol}, journal = {Biophysical Journal}, volume = {110}, number = {3, 1}, year = {2016}, note = {60th Annual Meeting of the Biophysical-Society, Los Angeles, CA, FEB 27-MAR 02, 2016}, month = {FEB}, pages = {356A-356A}, publisher = {Biophys Soc}, address = {600 TECHNOLOGY SQUARE, 5TH FLOOR, CAMBRIDGE, MA 02139 USA}, issn = {0006-3495}, author = {Sengupta, Durba and Prasanna, Xavier and Chattopadhyay, Amitabha} } @article {42992, title = {Ganglioside GM1 interacts with the serotonin1A receptor via the sphingolipid binding domain}, journal = {Biochimica et Biophysica Acta}, volume = {1858}, year = {2016}, month = {NOV}, pages = {2818-2826}, abstract = {

Glycosphingolipids are minor yet essential components of eukaryotic cell membranes and are involved in a variety of cellular processes. Although glycosphingolipids such as GM1 have been previously reported to influence the function of G protein-coupled receptors (GPCRs), the molecular mechanism remains elusive. In this paper, we have explored the interaction of GM1 with the serotonin1A receptor, an important neurotransmitter receptor that belongs to the GPCR family. To examine the molecular basis of the interaction of GM1 with the serotonin1A receptor, we performed a series of coarse-grain molecular dynamics simulations of the receptor embedded in membrane bilayers containing GM1. Our results show that GM1 interacts with the serotonin1A receptor predominantly at the extracellular loop 1 and specifically at the sphingolipid binding domain (SBD). The SBD motif consists of a characteristic combination of aromatic, basic and turn-inducing residues, and is evolutionarily conserved in case of the serotonin1A receptor. The interaction of the SBD site with GM1 appears to stabilize a {\textquoteright}flip-out{\textquoteright} conformation in which W102 of the extracellular loop 1 flips out from the central lumen of the receptor toward the membrane. The population of the {\textquoteright}flip-out{\textquoteright} conformation is increased in the presence of cholesterol. Our data strongly suggest that a direct interaction between GM1 and the SBD site of the serotonin1A receptor may occur in vivo. In view of the reported role of GM1 and the serotonin1A receptor in neurodegenerative diseases, GM1-receptor interaction assumes significance in the context of malfunctioning of neuronal GPCRs under such conditions.

}, doi = {10.1016/j.bbamem.2016.08.009}, author = {Prasanna, X. and Jafurulla, M. and Sengupta, Durba and Chattopadhyay, Amitabha} } @article {42993, title = {Protein-dependent membrane interaction of a partially disordered protein complex with oleic acid : Implications for cancer lipidomics}, journal = {Scientific Reports}, volume = {6}, year = {2016}, month = {OCT}, abstract = {

Bovine α-lactalbumin (BLA) forms cytotoxic complexes with oleic acid (OA) that perturbs tumor cell membranes, but molecular determinants of these membrane-interactions remain poorly understood. Here, we aim to obtain molecular insights into the interaction of BLA/BLA-OA complex with model membranes. We characterized the folding state of BLA-OA complex using tryptophan fluorescence and resolved residue-specific interactions of BLA with OA using molecular dynamics simulation. We integrated membrane-binding data using a voltage-sensitive probe and molecular dynamics (MD) to demonstrate the preferential interaction of the BLA-OA complex with negatively charged membranes. We identified amino acid residues of BLA and BLA-OA complex as determinants of these membrane interactions using MD, functionally corroborated by uptake of the corresponding α-LA peptides across tumor cell membranes. The results suggest that the α-LA component of these cytotoxic complexes confers specificity for tumor cell membranes through protein interactions that are maintained even in the lipid complex, in the presence of OA.

}, doi = {10.1038/srep35015}, author = {Chaudhuri, Arunima and Prasanna, Xavier and Agiru, Priyanka and Chakraborty, Hirak and Rydstrom, Anna and Ho, James C. S. and Svanborg, Catharina and Sengupta, Durba and Chattopadhyay, Amitabha} } @inbook { ISI:000375719300020, title = {What can simulations tell us about GPCRs: integrating the scales}, booktitle = {G Protein-Coupled Receptors: Signaling, Trafficking and Regulation}, series = {Methods in Cell Biology}, volume = {132}, year = {2016}, pages = {429-452}, publisher = {Elsevier Academic Press Inc.}, organization = {Elsevier Academic Press Inc.}, address = {525 B Street, Suite 1900, San Diego, CA 92101-4495 USA :}, abstract = {

The functional dynamics of G protein-coupled receptors (GPCRs) encompasses multiple spatiotemporal scales, ranging from femtoseconds to seconds and Angstroms to micrometers. Computational approaches, often in close collaboration with experimental methods, have been invaluable in unraveling GPCR structure and dynamics at these various hierarchical levels. The binding of natural and synthetic ligands to the wild-type and naturally occurring variant receptors have been analyzed by several computational methods. The activation of receptors from the inactive to the active state has been investigated by atomistic simulations and ongoing work on several receptors will help uncover general and receptor-specific mechanisms. The interaction of GPCRs with complex membranes that contain phospholipids and cholesterol have been probed by coarse-grain methods and shown to directly influence receptor association. In this chapter, we discuss computational approaches that have been successful in analyzing each scale of GPCR dynamics. An overview of these approaches will allow a more judicious choice of the appropriate method. We hope that an appreciation of the power of current computational approaches will encourage more critical collaborations. A comprehensive integration of the different approaches over the entire spatiotemporal scales promises to unravel new facets of GPCR function.

}, isbn = {978-0-12-803612-9; 978-0-12-803595-5}, issn = {0091-679X}, doi = {10.1016/bs.mcb.2015.11.007}, author = {Sengupta, Durba and Joshi, Manali and Athale, Chaitanya A. and Chattopadhyay, Amitabha}, editor = {Shukla, AK} } @article {44474, title = {Exploring GPCR lipid interactions by molecular dynamics simulations: excitements, challenges, and the way forward}, journal = {Journal of Physical Chemistry B}, volume = {122}, year = {2018}, month = {JUN}, pages = {5727-5737}, abstract = {Gprotein-coupled receptors (GPCRs) are seven transmembrane receptors that mediate a large number of cellular responses and are important drug targets. One of the current challenges in GPCR biology is to analyze the molecular signatures of receptor-lipid interactions and their subsequent effects on GPCR structure, organization, and function. Molecular dynamics simulation studies have been successful in predicting molecular determinants of receptor-lipid interactions. In particular, predicted cholesterol interaction sites appear to correspond well with experimentally determined binding sites and estimated time scales of association. In spite of several success stories, the methodologies in molecular dynamics simulations are still emerging. In this Feature Article, we provide a comprehensive overview of coarse-grain and atomistic molecular dynamics simulations of GPCR-lipid interaction in the context of experimental observations. In addition, we discuss the effect of secondary and tertiary structural constraints in coarse-grain simulations in the context of functional dynamics and structural plasticity of GPCRs. We envision that this comprehensive overview will help resolve differences in computational studies and provide a way forward.}, doi = {10.1021/acs.jpcb.8b01657}, author = {Sengupta, Durba and Prasanna, Xavier and Mohole, Madhura and Chattopadhyay, Amitabha} } @article {44521, title = {Membrane-induced organization and dynamics of the N-terminal domain of chemokine receptor CXCR1: insights from atomistic simulations}, journal = {Chemistry and Physics of Lipids}, volume = {210}, year = {2018}, month = {JAN}, pages = {142-148}, abstract = {

The CXC chemokine receptor 1 (CXCR1) is an important member of the G protein-coupled receptor (GPCR) family in which the extracellular N-terminal domain has been implicated in ligand binding and selectivity. The structure of this domain has not yet been elucidated due to its inherent dynamics, but experimental evidence points toward membrane-dependent organization and dynamics. To gain molecular insight into the interaction of the N-terminal domain with the membrane bilayer, we performed a series of microsecond time scale atomistic simulations of the N-terminal domain of CXCR1 in the presence and absence of POPC bilayers. Our results show that the peptide displays a high propensity to adopt a beta-sheet conformation in the presence of the membrane bilayer. The interaction of the peptide with the membrane bilayer was found to be transient in our simulations. Interestingly, a scrambled peptide, containing the same residues in a randomly varying sequence, did not exhibit membrane-modulated structural dynamics. These results suggest that sequence-dependent electrostatics, modulated by the membrane, could play an important role in folding of the N-terminal domain. We believe that our results reinforce the emerging paradigm that cellular membranes could be important modulators of function of G protein-coupled receptors such as CXCR1.

}, keywords = {All atom MD simulations, beta-sheet, CXCR1, Membrane interaction, N-terminal}, issn = {0009-3084}, doi = {10.1016/j.chemphyslip.2017.09.003}, author = {Kharche, Shalmali and Joshi, Manali and Sengupta, Durba and Chattopadhyay, Amitabha} } @inbook {47149, title = {Molecular determinants of GPCR oligomerization}, booktitle = {Molecular determinants of GPCR oligomerization}, number = {GPCRs}, year = {2020}, pages = {97-108}, doi = {10.1016/B978-0-12-816228-6.00006-4}, author = {Mohole, Madhura and Kumar, G. Aditya and Sengupta, Durba and Chattopadhyay, Amitabha} } @article {46943, title = {Role of cholesterol-mediated effects in GPCR heterodimers}, journal = {Chemistry and Physics of Lipids}, volume = {227}, year = {2020}, month = {MAR}, pages = {104852}, type = {Article}, abstract = {

G protein-coupled receptors (GPCRs) are transmembrane receptors that mediate a large number of cellular responses. The organization of GPCRs into dimers and higher-order oligomers is known to allow a larger repertoire of downstream signaling events. In this context, a crosstalk between the adenosine and dopamine receptors has been reported, indicating the presence of heterodimers that are functionally relevant. In this paper, we explored the effect of membrane cholesterol on the adenosine(2A) (A(2A)) and dopamine D-3 (D-3) receptors using coarse-grain molecular dynamics simulations. We analyzed cholesterol interaction sites on the A(2A) receptor and were able to reproduce the sites indicated by crystallography and previous atomistic simulations. We predict novel cholesterol interaction sites on the D-3 receptor that could be important in the reported cholesterol sensitivity in receptor function. Further, we analyzed the formation of heterodimers between the two receptors. Our results suggest that membrane cholesterol modulates the relative population of several co-existing heterodimer conformations. Both direct receptor-cholesterol interaction and indirect membrane effects contribute toward the modulation of heterodimer conformations. These results constitute one of the first examples of modulation of GPCR hetero-dimerization by membrane cholesterol, and could prove to be useful in designing better therapeutic strategies.

}, keywords = {Adenosine(2A) receptor, Cholesterol, Dopamine D-3 receptor, G protein-coupled receptors, Heterodimer, MARTINI coarse-grain simulation}, issn = {0009-3084}, doi = {10.1016/j.chemphyslip.2019.104852}, author = {Prasanna, Xavier and Mohole, Madhura and Chattopadhyay, Amitabha and Sengupta, Durba} } @article {47462, title = {Conformational plasticity and dynamic interactions of the N-terminal domain of the chemokine receptor CXCR1}, journal = {Plos Computational Biology}, volume = {17}, year = {2021}, month = {MAY}, type = {Article}, abstract = {

The dynamic interactions between G protein-coupled receptors (GPCRs) and their cognate protein partners are central to several cell signaling pathways. For example, the association of CXC chemokine receptor 1 (CXCR1) with its cognate chemokine, interleukin-8 (IL8 or CXCL8) initiates pathways leading to neutrophil-mediated immune responses. The N-terminal domain of chemokine receptors confers ligand selectivity, but unfortunately the conformational dynamics of this intrinsically disordered region remains unresolved. In this work, we have explored the interaction of CXCR1 with IL8 by microsecond time scale coarse-grain simulations, complemented by atomistic models and NMR chemical shift predictions. We show that the conformational plasticity of the apo-receptor N-terminal domain is restricted upon ligand binding, driving it to an open C-shaped conformation. Importantly, we corroborated the dynamic complex sampled in our simulations against chemical shift perturbations reported by previous NMR studies and show that the trends are similar. Our results indicate that chemical shift perturbation is often not a reporter of residue contacts in such dynamic associations. We believe our results represent a step forward in devising a strategy to understand intrinsically disordered regions in GPCRs and how they acquire functionally important conformational ensembles in dynamic protein-protein interfaces.

}, doi = {10.1371/journal.pcbi.1008593}, author = {Kharche, Shalmali and Joshi, Manali and Chattopadhyay, Amitabha and Sengupta, Durba} } @article {48347, title = {Synergistic and competitive lipid interactions in the serotonin1A receptor microenvironment}, journal = {ACS Chemical Neuroscience}, volume = {13}, year = {2022}, month = {DEC }, pages = {3403-3415}, type = {Article}, abstract = {

The interaction of lipids with G-protein-coupled receptors (GPCRs) has been shown to modulate and dictate several aspects of GPCR organization and function. Diverse lipid interaction sites have been identified from structural biology, bioinformatics, and molecular dynamics studies. For example, multiple cholesterol interaction sites have been identified in the serotonin1A receptor, along with distinct and overlapping sphingolipid interaction sites. How these lipids interact with each other and what is the resultant effect on the receptor is still not clear. In this work, we have analyzed lipid-lipid crosstalk at the receptor of the serotonin1A receptor embedded in a membrane bilayer that mimics the neuronal membrane composition by long coarse-grain simulations. Using a set of similarity coefficients, we classified lipids that bind at the receptor together as synergistic cobinding, and those that bind individually as competitive. Our results show that certain lipids interact with the serotonin1A receptor in synergy with each other. Not surprisingly, the ganglioside GM1 and cholesterol show a synergistic cobinding, along with the relatively uncommon GM1-phosphatidylethanolamine (PE) and cholesterol-PE synergy. In contrast, certain lipid pairs such as cholesterol and sphingomyelin appear to be in competition at several sites, despite their coexistence in lipid nanodomains. In addition, we observed intralipid competition between two lipid tails, with the receptor exhibiting increased interactions with the unsaturated lipid tails. We believe our work represents an important step in understanding the diversity of GPCR-lipid interactions and exploring synergistic cobinding and competition in natural membranes.

}, keywords = {GPCR, lipid crosstalk, Martini coarse-grain simulations, Membrane cholesterol, neuronal membranes}, issn = {1948-7193}, doi = {10.1021/acschemneuro.2c00422}, author = {Mohole, Madhura and Sengupta, Durba and Chattopadhyay, Amitabha} }