01702nas a2200157 4500008004100000245007200041210006900113260000800182300001200190490000700202520116900209100002401378700002101402700002001423856010101443 2015 eng d00aEffect of lipid bilayer composition on membrane protein association0 aEffect of lipid bilayer composition on membrane protein associat cJUL a43–630 v223 a

Diverse cellular functions are supported by membrane protein assemblies associated with the cell membrane. Although considered to be protein-mediated, membrane components are now being recognized as critical in modulating and sometime dictating function. This chapter discusses the effect of the lipid bilayer, in particular its composition on membrane protein organization. Computational methods have been successful in quantifying transmembrane protein association and general features of dimerization profiles are explored. Understanding the molecular basis of the interactions has lead to the recognition of the lipophobic effects. These nonspecific effects include those that arise from membrane perturbations and lipid chain packing and have been shown to modulate the energetics as well as the structural characteristics of membrane protein dimerization. In addition, specific interactions arising from direct protein–lipid interactions and protein–cholesterol interactions have been suggested to influence membrane protein association. We summarize here a few examples highlighting the role of the lipid bilayer on membrane protein organization.

1 aPawar, Aiswarya, B.1 aPrasanna, Xavier1 aSengupta, Durba uhttps://library.ncl.res.in/content/effect-lipid-bilayer-composition-membrane-protein-association02001nas a2200205 4500008004100000022001400041245008200055210006900137260011000206300001400316490000700330520119600337100002401533700002501557700002501582700002601607700002601633700002001659856011601679 2015 eng d a1463-907600aThermodynamic and kinetic characterization of transmembrane helix association0 aThermodynamic and kinetic characterization of transmembrane heli aTHOMAS GRAHAM HOUSE, SCIENCE PARK, MILTON RD, CAMBRIDGE CB4 0WF, CAMBS, ENGLANDbROYAL SOC CHEMISTRYcNOV a1390-13980 v173 a

The transient dimerization of transmembrane proteins is an important event in several cellular processes and computational methods are being increasingly used to quantify their underlying energetics. Here, we probe the thermodynamics and kinetics of a simple transmembrane dimer to understand membrane protein association. A multi-step framework has been developed in which the dimerization profiles are calculated from coarse-grain molecular dynamics simulations, followed by meso-scale simulations using parameters calculated from the coarse-grain model. The calculated value of Delta G(assoc) is approx. -20 kJ mol(-1) and is consistent between three methods. Interestingly, the meso-scale stochastic model reveals low dimer percentages at physiologically-relevant concentrations, despite a favorable Delta G(assoc). We identify generic driving forces arising from the protein backbone and lipid bilayer and complementary factors, such as protein density, that govern self-interactions in membranes. Our results provide an important contribution in understanding membrane protein organization and linking molecular, nano-scale computational studies to meso-scale experimental data.

1 aPawar, Aiswarya, B.1 aDeshpande, Sneha, A.1 aGopal, Srinivasa, M.1 aWassenaar, Tsjerk, A.1 aAthale, Chaitanya, A.1 aSengupta, Durba uhttps://library.ncl.res.in/content/thermodynamic-and-kinetic-characterization-transmembrane-helix-association-002400nas a2200181 4500008004100000245010500041210006900146260000800215300001100223490000900234520173200243100002501975700002402000700001702024700002602041700002002067856013102087 2017 eng d00aRole of spatial inhomogenity in GPCR dimerisation predicted by receptor association-diffusion models0 aRole of spatial inhomogenity in GPCR dimerisation predicted by r cJUN a0360020 v14 3 aG protein-coupled receptor (GPCR) association is an emerging paradigm with far reaching implications in the regulation of signalling pathways and therapeutic interventions. Recent super resolution microscopy studies have revealed that receptor dimer steady state exhibits sub-second dynamics. In particular the GPCRs, muscarinic acetylcholine receptor M-1 (M1MR) and formyl peptide receptor (FPR), have been demonstrated to exhibit a fast association/dissociation kinetics, independent of ligand binding. In this work, we have developed a spatial kinetic Monte Carlo model to investigate receptor homo-dimerisation at a single receptor resolution. Experimentally measured association/dissociation kinetic parameters and diffusion coefficients were used as inputs to the model. To test the effect of membrane spatial heterogeneity on the simulated steady state, simulations were compared to experimental statistics of dimerisation. In the simplest case the receptors are assumed to be diffusing in a spatially homogeneous environment, while spatial heterogeneity is modelled to result from crowding, membrane micro-domains and cytoskeletal compartmentalisation or 'corrals'. We show that a simple association-diffusion model is sufficient to reproduce M1MR association statistics, but fails to reproduce FPR statistics despite comparable kinetic constants. A parameter sensitivity analysis is required to reproduce the association statistics of FPR. The model reveals the complex interplay between cytoskeletal components and their influence on receptor association kinetics within the features of the membrane landscape. These results constitute an important step towards understanding the factors modulating GPCR organisation.1 aDeshpande, Sneha, A.1 aPawar, Aiswarya, B.1 aDighe, Anish1 aAthale, Chaitanya, A.1 aSengupta, Durba uhttps://library.ncl.res.in/content/role-spatial-inhomogenity-gpcr-dimerisation-predicted-receptor-association-diffusion-models01941nas a2200193 4500008004100000022001400041245008000055210006900135260000900204300001200213490000800225520126800233653003901501653003101540653002401571100002401595700002001619856010801639 2019 eng d a1047-847700aResolving the conformational dynamics of ErbB growth factor receptor dimers0 aResolving the conformational dynamics of ErbB growth factor rece cAUG  a225-2330 v2073 a

The combinatorial dimerization of the ErbB growth factor receptors (ErbB(1) - ErbB(4)) are critical for their function. Here, we have characterized the conformational dynamics of ErbB transmembrane homo-dimers and hetero-dimers by using a coarse-grain simulation framework. All dimers, except ErbB (4-4) and ErbB(1-4), exhibit at least two conformations. The reported NMR structures correspond to one of these conformations, representing the N-terminal active state in ErbB(1-1) (RH2), ErbB(2-2) (RH1) and ErbB(4-4) (RH) homo-dimers and the LH dimer in ErbB(3-3) homo-dimer, validating the computational approach. Further, we predict a right-handed ErbB(4) dimer conformer that warrants experimental testing. The five hetero-dimers that have not yet been experimentally resolved display prominent right-handed dimers associating by the SmXXXSm motif. Our results provide insights into the constitutive signaling of ErbB(4) after cleavage of the extracellular region. The presence of the inactive-like dimer conformers leading to symmetric kinase domains gives clues on the autoinhibition of the receptor dimers. The dimer states characterized here represent an important step towards understanding the combinatorial cross associations in the ErbB family.

10aMartini coarse-grained simulations10aMembrane protein structure10aTransmembrane dimer1 aPawar, Aiswarya, B.1 aSengupta, Durba uhttps://library.ncl.res.in/content/resolving-conformational-dynamics-erbb-growth-factor-receptor-dimers02311nas a2200241 4500008004100000022001400041245008800055210006900143260000800212300001100220490000800231520148500239653001601724653001801740653003901758653003101797653003001828100002501858700002401883700002601907700002001933856011601953 2020 eng d a0009-308400aLipid-dependent conformational landscape of the ErbB2 growth factor receptor dimers0 aLipiddependent conformational landscape of the ErbB2 growth fact cAUG a1049110 v2303 a

Altered lipid metabolism has been linked to cancer development and progression. Several roles have been attributed to the increased saturation and length of lipid acyl tails observed in tumors, but its effect on signaling receptors is still emerging. In this work, we have analyzed the lipid dependence of the ErbB2 growth factor receptor dimerization that plays an important role in the pathogenesis of breast cancer. We have performed coarse-grain ensemble molecular dynamics simulations to comprehensively sample the ErbB2 monomer-dimer association. Our results indicate a dynamic dimer state with a complex conformational landscape that is modulated with increasing lipid tail length. We resolve the native N-terminal ``active'' and C-terminal ``inactive'' conformations in all membrane compositions. However, the relative population of the N-terminal and C-terminal conformers is dependent on length of the saturated lipid tails. In short-tail membranes, additional non-specific dimers are observed which are reduced or absent in long-tailed bilayers. Our results indicate that the relative population as well as the structure of the dimer state is modulated by membrane composition. We have correlated these differences to local perturbations of the membrane around the receptor. Our work is an important step in characterizing ErbB dimers in healthy and diseased states and emphasize the importance of sampling lipid dynamics in understanding receptor association.

10aErbB2 dimer10aLipid effects10aMartini coarse-grained simulations10aMembrane protein structure10aTransmembrane association1 aGopal, Srinivasa, M.1 aPawar, Aiswarya, B.1 aWassenaar, Tsjerk, A.1 aSengupta, Durba uhttps://library.ncl.res.in/content/lipid-dependent-conformational-landscape-erbb2-growth-factor-receptor-dimers02054nas a2200205 4500008004100000022001400041245009000055210006900145260000800214520132800222653001601550653003301566653001501599653002401614653003701638653001701675100002401692700002001716856011201736 2021 eng d a0022-263100aRole of cholesterol in transmembrane dimerization of the ErbB2 growth factor receptor0 aRole of cholesterol in transmembrane dimerization of the ErbB2 g cJAN3 a

The association of ErbB2 growth factor receptors is critical for cell growth and potentiates tumor proliferation in several cancer types. An important aspect in ErbB2 association is the role of lipids such as cholesterol, especially since their metabolism is often reprogrammed in cancer cells. Here, we have coupled metadynamics with coarse-grain simulations to identify cholesterol effects in the transmembrane dimerization of ErbB2 receptors. Overall, cholesterol interactions are observed with the receptor that directly tunes the association energetics. Several dimer conformations are identified both in the presence and absence of cholesterol, although the dimer regime appears to be more favorable in the presence of cholesterol. We observe an overall modulation of the underlying energy profile and the symmetric active and inactive conformational states are not distinguished in the presence of cholesterol. We show that cholesterol binds to the receptor transmembrane domain at a site (CRAC motif) that overlaps with the dimer interface (SmXXXSm motif). The competition between the transmembrane interactions and cholesterol interactions decides the final conformational landscape. Our work is an important step toward characterizing cholesterol effects in ErbB2 membrane receptor function. [GRAPHICS] .

10aCholesterol10aErbB2 growth factor receptor10aHer2 dimer10aJuxtamembrane dimer10aMartini coarse-grain simulations10aMetadynamics1 aPawar, Aiswarya, B.1 aSengupta, Durba uhttps://library.ncl.res.in/content/role-cholesterol-transmembrane-dimerization-erbb2-growth-factor-receptor02685nas a2200229 4500008004100000022001400041245016600055210006900221260000800290300001600298490000600314520183500320100001802155700002402173700002002197700002302217700002002240700002002260700002002280700002502300856013002325 2022 eng d a2470-134300aInvestigation of the captopril-insulin interaction by mass spectrometry and computational approaches reveals that captopril induces structural changes in insulin0 aInvestigation of the captoprilinsulin interaction by mass spectr cJUL a23115-231260 v73 a

ABSTRACT: Post-translational modifications remarkably regulate proteins' biological function. Small molecules such as reactive thiols, metabolites, and drugs may covalently modify the proteins and cause structural changes. This study reports the covalent modification and noncovalent interaction of insulin and captopril, an FDA-approved antihypertensive drug, through mass spectrometric and computation-based approaches. Mass spectrometric analysis shows that captopril modifies intact insulin, reduces it into its ``A'' and ``B'' chains, and covalently modifies them by forming adducts. Since captopril has a reactive thiol group, it might reduce the insulin dimer or modify it by reacting with cysteine residues. This was proven with dithiothreitol treatment, which reduced the abundance of captopril adducts of insulin A and B chains and intact Insulin. Liquid chromatography tandem mass spectrometric analysis identified the modification of a total of four cysteine residues, two in each of the A and B chains of insulin. These modifications were identified to be Cys6 and Cys7 of the A chain and Cys7 and Cys19 of the B chain. Mass spectrometric analysis indicated that captopril may simultaneously modify the cysteine residues of intact insulin or its subunits A and B chains. Biophysical studies involving light scattering and thioflavin T assay suggested that the binding of captopril to the protein leads to the formation of aggregates. Docking and molecular dynamics studies provided insights into the noncovalent interactions and associated structural changes in insulin. This work is a maiden attempt to understand the detailed molecular interactions between captopril and insulin. These findings suggest that further investigations are required to understand the long-term effect of drugs like captopril.

1 aGhosh, Amrita1 aPawar, Aiswarya, B.1 aChirmade, Tejas1 aJathar, Swaraj, M.1 aBhambure, Rahul1 aSengupta, Durba1 aGiri, Ashok, P.1 aKulkarni, Mahesh, J. uhttps://library.ncl.res.in/content/investigation-captopril-insulin-interaction-mass-spectrometry-and-computational-approaches