Costing of a Combination Intervention (Kyaterekera) Addressing Sexual Risk-Taking Behaviors among Vulnerable Women in Southern Uganda.

In Uganda, women engaged in sex work (WESW) are a marginalized population at the intersection of multiple vulnerabilities. The Kyaterekera intervention is targeted at WESW in Rakai and the greater Masaka regions in Uganda and combines a traditional HIV risk-reduction approach with a savings-led economic empowerment intervention and financial literacy training. We estimated the economic costs of the Kyaterekera intervention from a program provider perspective using a prospective activity-based micro-costing method. All program activities and resource uses were measured and valued across the control arm receiving a traditional HIV risk-reduction intervention and the treatment arm receiving a matched individual development savings account and financial literacy training on top of HIV risk reduction. The total per-participant cost by arm was adjusted for inflation and discounted at an annual rate of 3% and presented in 2019 US dollars. The total per-participant costs of the control and intervention arms were estimated at $323 and $1,435, respectively, using the per-protocol sample. When calculated based on the intent-to-treat sample, the per-participant costs were reduced to $183 and $588, respectively. The key cost drivers were the capital invested in individual development accounts and personnel and transportation costs for program operations, linked to WESW's higher mobility and the dispersed pattern of hot spot locations. The findings provide evidence of the economic costs of implementing a targeted intervention for this marginalized population in resource-constrained settings and shed light on the scale of potential investment needed to better achieve the health equity goal of HIV prevention strategies.

Dual activity of Minnelide chemosensitize basal/triple negative breast cancer stem cells and reprograms immunosuppressive tumor microenvironment.

Triple negative breast cancer (TNBC) subtype is characterized with higher EMT/stemness properties and immune suppressive tumor microenvironment (TME). Women with advanced TNBC exhibit aggressive disease and have limited treatment options. Although immune suppressive TME is implicated in driving aggressive properties of basal/TNBC subtype and therapy resistance, effectively targeting it remains a challenge. Minnelide, a prodrug of triptolide currently being tested in clinical trials, has shown anti-tumorigenic activity in multiple malignancies via targeting super enhancers, Myc and anti-apoptotic pathways such as HSP70. Distinct super-enhancer landscape drives cancer stem cells (CSC) in TNBC subtype while inducing immune suppressive TME. We show that Minnelide selectively targets CSCs in human and murine TNBC cell lines compared to cell lines of luminal subtype by targeting Myc and HSP70. Minnelide in combination with cyclophosphamide significantly reduces the tumor growth and eliminates metastasis by reprogramming the tumor microenvironment and enhancing cytotoxic T cell infiltration in 4T1 tumor-bearing mice. Resection of residual tumors following the combination treatment leads to complete eradication of disseminated tumor cells as all mice are free of local and distant recurrences. All control mice showed recurrences within 3 weeks of post-resection while single Minnelide treatment delayed recurrence and one mouse was free of tumor. We provide evidence that Minnelide targets tumor intrinsic pathways and reprograms the immune suppressive microenvironment. Our studies also suggest that Minnelide in combination with cyclophosphamide may lead to durable responses in patients with basal/TNBC subtype warranting its clinical investigation.

Synthesis, Structural Investigations, DNA/BSA Interactions, Molecular Docking Studies, and Anticancer Activity of a New 1,4-Disubstituted 1,2,3-Triazole Derivative.

We report herein a new 1,2,3-triazole derivative, namely, 4-((1-(3,4-dichlorophenyl)-1H-1,2,3-triazol-4-yl)methoxy)-2-hydroxybenzaldehyde, which was synthesized by copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC). The structure of the compound was analyzed using Fourier transform infrared spectroscopy (FTIR), 1H NMR, 13C NMR, UV-vis, and elemental analyses. Moreover, X-ray crystallography studies demonstrated that the compound adapted a monoclinic crystal system with the P21/c space group. The dominant interactions formed in the crystal packing were found to be hydrogen bonding and van der Waals interactions according to Hirshfeld surface (HS) analysis. The volume of the crystal voids and the percentage of free spaces in the unit cell were calculated as 152.10 Å3 and 9.80%, respectively. The evaluation of energy frameworks showed that stabilization of the compound was dominated by dispersion energy contributions. Both in vitro and in silico investigations on the DNA/bovine serum albumin (BSA) binding activity of the compound showed that the CT-DNA binding activity of the compound was mediated via intercalation and BSA binding activity was mediated via both polar and hydrophobic interactions. The anticancer activity of the compound was also tested by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay using human cell lines including MDA-MB-231, LNCaP, Caco-2, and HEK-293. The compound exhibited more cytotoxic activity than cisplatin and etoposide on Caco-2 cancer cell lines with an IC50 value of 16.63 ± 0.27 µM after 48 h. Annexin V suggests the induction of cell death by apoptosis. Compound 3 significantly increased the loss of mitochondrial membrane potential (MMP) levels in Caco-2 cells, and the reactive oxygen species (ROS) assay proved that compound 3 could induce apoptosis by ROS generation.

Nonlocal Effects of Antibiotic-Resistance-Causing Mutations Reveal an Alternative Region for Targeting on FtsW-Penicillin-Binding Protein 3 Complex of Haemophilus influenzae.

Currently prescribed antibiotics target the catalytic sites of wild-type bacterial proteins; however, bacteria adopt mutations at this site, eventually leading to the emergence of resistance. Therefore, the identification of alternative drug binding sites is crucial, which requires knowledge of the dynamics of the mutant protein. Here, we set out to investigate the impact of a high-resistance-causing triple mutation (S385T + L389F + N526K) on the dynamics of a prioritized resistant pathogen, Haemophilus influenzae, using computational techniques. We studied penicillin-binding protein 3 (PBP3) and its complex with FtsW, which display resistance toward ß-lactam antibiotics. We showed that mutations displayed local and nonlocal effects. In terms of the former, the orientation of the ß-sheet, which surrounds the active site of PBP3, was impacted and the catalytic site was exposed to the periplasmic region. In addition, the flexibility of the ß3-ß4 loop, which modulates the catalysis of the enzyme, increased in the mutant FtsW-PBP3 complex. As for nonlocal effects, the dynamics of the pedestal domain (N-terminal periplasmic modulus (N-t)), i.e., the opening of the fork, was different between the wild-type and mutant enzymes. We showed the closed fork caused a greater number of residues to participate in the hypothesized allosteric communication network connecting N-t to the transpeptidase domain in the mutant enzyme. Finally, we demonstrated that the closed fork results in more favorable binding with ß-lactam antibiotics, particularly cefixime, suggesting that small therapeutics that can stabilize the closed fork of mutant PBP3 may lead to the development of more effective molecules to combat resistant bacteria.

Inhibition of mutant RAS-RAF interaction by mimicking structural and dynamic properties of phosphorylated RAS.

Undruggability of RAS proteins has necessitated alternative strategies for the development of effective inhibitors. In this respect, phosphorylation has recently come into prominence as this reversible post-translational modification attenuates sensitivity of RAS towards RAF. As such, in this study, we set out to unveil the impact of phosphorylation on dynamics of HRASWT and aim to invoke similar behavior in HRASG12D mutant by means of small therapeutic molecules. To this end, we performed molecular dynamics (MD) simulations using phosphorylated HRAS and showed that phosphorylation of Y32 distorted Switch I, hence the RAS/RAF interface. Consequently, we targeted Switch I in HRASG12D by means of approved therapeutic molecules and showed that the ligands enabled detachment of Switch I from the nucleotide-binding pocket. Moreover, we demonstrated that displacement of Switch I from the nucleotide-binding pocket was energetically more favorable in the presence of the ligand. Importantly, we verified computational findings in vitro where HRASG12D/RAF interaction was prevented by the ligand in HEK293T cells that expressed HRASG12D mutant protein. Therefore, these findings suggest that targeting Switch I, hence making Y32 accessible might open up new avenues in future drug discovery strategies that target mutant RAS proteins.

Mechanistic insight into impact of phosphorylation on the enzymatic steps of farnesyltransferase.

Farnesyltransferase (FTase) is a heterodimeric enzyme, which catalyzes covalent attachment of the farnesyl group to target proteins, thus coordinating their trafficking in the cell. FTase has been demonstrated to be highly expressed in cancer and neurological diseases; hence considered as a hot target for therapeutic purposes. However, due to the nonspecific inhibition, there has been only one inhibitor that could be translated into the clinic. Importantly, it has been shown that phosphorylation of the α-subunit of FTase increases the activity of the enzyme in certain diseases. As such, understanding the impact of phosphorylation on dynamics of FTase provides a basis for targeting a specific state of the enzyme that emerges under pathological conditions. To this end, we performed 18 µs molecular dynamics (MD) simulations using complexes of (non)-phosphorylated FTase that are representatives of the farnesylation reaction. We demonstrated that phosphorylation modulated the catalytic site by rearranging interactions between farnesyl pyrophosphate (FPP)/peptide substrate, catalytic Zn2+ ion/coordinating residues and hot-spot residues at the interface of the subunits, all of which led to the stabilization of the substrate and facilitation of the release of the product, thus collectively expediting the reaction rate. Importantly, we also identified a likely allosteric pocket on the phosphorylated FTase, which might be used for specific targeting of the enzyme. To the best of our knowledge, this is the first study that systematically examines the impact of phosphorylation on the enzymatic reaction steps, hence opens up new avenues for drug discovery studies that focus on targeting phosphorylated FTase.

Experimental and Computational Investigation of Clustering Behavior of Cyclodextrin-Perfluorocarbon Inclusion Complexes as Effective Histotripsy Agents.

Recently developed nanocones (NCs), which are inclusion complexes that are made up of cyclodextrins (CDs) and perfluorocarbons (PFCs), have shown promising results in nanoparticle-mediated histotripsy (NMH) applications due to stable inclusion complexation, PFC quantification, simple synthesis, and processing. FDA-approved ßCD and its modified versions such as low-degree methylated ßCD have been previously demonstrated as prime examples of structures capable of accommodating PFC molecules. However, the complex formation potential of different CDs with various cavity sizes in the presence of PFC molecules, and their consequent aggregation, needs to be explored. In the present study, the complexation and aggregation potential of some natural CDs and their respective derivatives either exposed to perfluoropentane (PFP) or perfluorohexane (PFH) were studied in the wet lab. Computational studies were also performed to account for the limitations faced in PFC quantification because of the low optical density of PFCs within the CD complex and to discover the best candidate for NMH applications. All results revealed that only ßCD and γCD (except HMγCD) derivatives form an inclusion complex with PFCs and only LMßCD, ßCD, and γCD form nanocone clusters (NCCs), which precipitate and can be collected for use. Furthermore, the data collectively show that ßCD and PFCs have the best complexation due to stable complex formation, ease of production, and product recovery, especially with PFH as a more suitable candidate due to its high boiling point, which allows workability during synthesis. Although simulations suggest that highly stable inclusion complexes exist, such as HPßCD, the cluster formation resulting in precipitation is hindered due to the high solubility of CDs in water, resulting in intangible yields to work with even after employing general laboratory recovery methods. Conclusively, histotripsy cavitation experiments successfully showed a decreased cavitation threshold among optimal NCC candidates that were identified, supporting their use in NMH.

Mechanistic insight into the impact of a bivalent ligand on the structure and dynamics of a GPCR oligomer.

Development of effective bivalent ligands has become the focus of intensive research toward modulation of G protein-coupled receptor (GPCR) oligomers, particularly in the field of GPCR pharmacology. Experimental studies have shown that they increased binding affinity and signaling potency compared to their monovalent counterparts, yet underlying molecular mechanism remains elusive. To address this, we performed accelerated molecular dynamics simulations on bivalent-ligand bound Adenosine 2A receptor (A2AR) dimer in the context of a modeled tetramer, which consists of A2AR and dopamine 2 receptor (D2R) homodimers and their cognate G proteins. Our results demonstrate that bivalent ligand impacted interactions between pharmacophore groups and ligand binding residues, thus modulating allosteric communication network and water channel formed within the receptor. Moreover, it also strengthens contacts between receptor and G protein, by modulating the volume of ligand binding pocket and intracellular domain of the receptor. Importantly, we showed that impact evoked by the bivalent ligand on A2AR dimer was also transmitted to apo D2R, which is part of the neighboring D2R dimer. To the best of our knowledge, this is the first study that provides a mechanistic insight into the impact of a bivalent ligand on dynamics of a GPCR oligomer. Consequently, this will pave the way for development of effective ligands for modulation of GPCR oligomers and hence treatment of crucial diseases such as Parkinson's disease and cancer.

Inhibition of SARS-CoV-2 main protease: a repurposing study that targets the dimer interface of the protein.

Coronavirus disease-2019 (COVID-19) was firstly reported in Wuhan, China, towards the end of 2019, and emerged as a pandemic. The spread and lethality rates of the COVID-19 have ignited studies that focus on the development of therapeutics for either treatment or prophylaxis purposes. In parallel, drug repurposing studies have also come into prominence. Herein, we aimed at having a holistic understanding of conformational and dynamical changes induced by an experimentally characterized inhibitor on main protease (Mpro) which would enable the discovery of novel inhibitors. To this end, we performed molecular dynamics simulations using crystal structures of apo and α-ketoamide 13b-bound Mpro homodimer. Analysis of trajectories pertaining to apo Mpro revealed a new target site, which is located at the homodimer interface, next to the catalytic dyad. Thereafter, we performed ensemble-based virtual screening by exploiting the ZINC and DrugBank databases and identified three candidate molecules, namely eluxadoline, diosmin, and ZINC02948810 that could invoke local and global conformational rearrangements which were also elicited by α-ketoamide 13b on the catalytic dyad of Mpro. Furthermore, ZINC23881687 stably interacted with catalytically important residues Glu166 and Ser1 and the target site throughout the simulation. However, it gave positive binding energy, presumably, due to displaying higher flexibility that might dominate the entropic term, which is not included in the MM-PBSA method. Finally, ZINC20425029, whose mode of action was different, modulated dynamical properties of catalytically important residue, Ala285. As such, this study presents valuable findings that might be used in the development of novel therapeutics against Mpro.Communicated by Ramaswamy H. Sarma.

Dimethyl sulfoxide reduces the stability but enhances catalytic activity of the main SARS-CoV-2 protease 3CLpro.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for coronavirus disease 2019 (COVID-19), one of the most challenging global pandemics of the modern era. Potential treatment strategies against COVID-19 are yet to be devised. It is crucial that antivirals that interfere with the SARS-CoV-2 life cycle be identified and developed. 3-Chymotrypsin-like protease (3CLpro) is an attractive antiviral drug target against SARS-CoV-2, and coronaviruses in general, because of its role in the processing of viral polyproteins. Inhibitors of 3CLpro activity are screened in enzyme assays before further development of the most promising leads. Dimethyl sulfoxide (DMSO) is a common additive used in such assays and enhances the solubility of assay components. However, it may also potentially affect the stability and efficiency of 3CLpro but, to date, this effect had not been analyzed in detail. Here, we investigated the effect of DMSO on 3CLpro-catalyzed reaction. While DMSO (5%-20%) decreased the optimum temperature of catalysis and thermodynamic stability of 3CLpro, it only marginally affected the kinetic stability of the enzyme. Increasing the DMSO concentration up to 20% improved the catalytic efficiency and peptide-binding affinity of 3CLpro. At such high DMSO concentration, the solubility and stability of peptide substrate were improved because of reduced aggregation. In conclusion, we recommend 20% DMSO as the minimum concentration to be used in screens of 3CLpro inhibitors as lead compounds for the development of antiviral drugs against COVID-19.

Linker residues regulate the activity and stability of hexokinase 2, a promising anticancer target.

Hexokinase (HK) catalyzes the first step in glucose metabolism, making it an exciting target for the inhibition of tumor initiation and progression due to their elevated glucose metabolism. The upregulation of hexokinase-2 (HK2) in many cancers and its limited expression in normal tissues make it a particularly attractive target for the selective inhibition of cancer growth and the eradication of tumors with limited side effects. The design of such safe and effective anticancer therapeutics requires the development of HK2-specific inhibitors that will not interfere with other HK isozymes. As HK2 is unique among HKs in having a catalytically active N-terminal domain (NTD), we have focused our attention on this region. We previously found that NTD activity is affected by the size of the linker helix-α13 that connects the N- and C-terminal domains of HK2. Three nonactive site residues (D447, S449, and K451) at the beginning of the linker helix-α13 have been found to regulate the NTD activity of HK2. Mutation of these residues led to increased dynamics, as shown via hydrogen deuterium exchange analysis and molecular dynamic simulations. D447A contributed the most to the enhanced dynamics of the NTD, with reduced calorimetric enthalpy of HK2. Similar residues exist in the C-terminal domain (CTD) but are unnecessary for HK1 and HK2 activity. Thus, we postulate these residues serve as a regulatory site for HK2 and may provide new directions for the design of anticancer therapeutics that reduce the rate of glycolysis in cancer through specific inhibition of HK2.

In Silico Analysis of a Highly Mutated Gene in Cancer Provides Insight into Abnormal mRNA Splicing: Splicing Factor 3B Subunit 1K700E Mutant.

Cancer is the second leading cause of death worldwide. The etiology of the disease has remained elusive, but mutations causing aberrant RNA splicing have been considered one of the significant factors in various cancer types. The association of aberrant RNA splicing with drug/therapy resistance further increases the importance of these mutations. In this work, the impact of the splicing factor 3B subunit 1 (SF3B1) K700E mutation, a highly prevalent mutation in various cancer types, is investigated through molecular dynamics simulations. Based on our results, K700E mutation increases flexibility of the mutant SF3B1. Consequently, this mutation leads to i) disruption of interaction of pre-mRNA with SF3B1 and p14, thus preventing proper alignment of mRNA and causing usage of abnormal 3' splice site, and ii) disruption of communication in critical regions participating in interactions with other proteins in pre-mRNA splicing machinery. We anticipate that this study enhances our understanding of the mechanism of functional abnormalities associated with splicing machinery, thereby, increasing possibility for designing effective therapies to combat cancer at an earlier stage.

Perturb-Scan-Pull: A Novel Method Facilitating Conformational Transitions in Proteins.

Conformational transitions in proteins facilitate precise physiological functions. Therefore, it is crucial to understand the mechanisms underlying these processes to modulate protein function. Yet, studying structural and dynamical properties of proteins is notoriously challenging due to the complexity of the underlying potential energy surfaces (PES). We have previously developed the perturbation-response scanning (PRS) method to identify key residues that participate in the communication network responsible for specific conformational transitions. PRS is based on a residue-by-residue scan of the protein to determine the subset of residues/forces which provide the closest conformational change leading to a target conformational state, inasmuch as linear response theory applies to these motions. Here, we develop a novel method to further evaluate if conformational transitions may be triggered on the PES. We aim to study functionally relevant conformational transitions in proteins by using results obtained from PRS and feeding them as inputs to steered molecular dynamics simulations. The success and the transferability of the method are evaluated on three protein systems having different complexities of motion on the PES: calmodulin, adenylate kinase, and bacterial ferric binding protein. We find that the method captures the target conformation, while providing key residues and the optimum paths with relatively low free energy profiles.

The single nucleotide ß -arrestin2 variant, A248T, resembles dynamical properties of activated arrestin.

ß -arrestins are responsible for termination of G protein-coupled receptor (GPCR)-mediated signaling. Association of single nucleotide variants with onset of crucial diseases has made this protein family hot targets in the field of GPCR-mediated pharmacology. However, impact of these mutations on function of these variants has remained elusive. In this study, structural and dynamical properties of one of ß -arrestin2 (arrestin 3) variants, A248T, which has been identified in some cancer tissue samples, were investigated via molecular dynamics simulations. The results showed that the variant underwent structural rearrangements which are seen in crystal structures of active arrestin. Specifically, the "short helix" unravels and the "gate loop" swings forward as seen in crystal structures of receptor-bound and GPCR phosphopeptide-bound arrestin. Moreover, the "finger loop" samples upward position in the variant. Importantly, these regions harbor crucial residues that are involved in receptor binding interfaces. Cumulatively, these local structural rearrangements help the variant adopt active-like domain angle without perturbing the "polar core". Considering that phosphorylation of the receptor is required for activation of arrestin, A248T might serve as a model system to understand phosphorylation-independent activation mechanism, thus enabling modulation of function of arrestin variants which are activated independent of receptor phosphorylation as seen in cancer.

Catalytically Competent Non-transforming H-RASG12P Mutant Provides Insight into Molecular Switch Function and GAP-independent GTPase Activity of RAS.

RAS mutants have been extensively studied as they are associated with development of cancer; however, H-RASG12P mutant has remained untouched since it does not lead to transformation in the cell. To the best of our knowledge, this is the first study where structural/dynamical properties of H-RASG12P have been investigated -in comparison to H-RASWT, H-RASG12D, RAF-RBD-bound and GAP-bound H-RASWT- using molecular dynamics simulations (total of 9 µs). We observed remarkable differences in dynamics of Y32. Specifically, it is located far from the nucleotide binding pocket in the catalytically-active GAP-bound H-RASWT, whereas it makes close interaction with the nucleotide in signaling-active systems (H-RASG12D, KRAS4BG12D, RAF-RBD-bound H-RASWT) and H-RASWT. The accessibility of Y32 in wild type protein is achieved upon GAP binding. Interestingly; however, it is intrinsically accessible in H-RASG12P. Considering the fact that incomplete opening of Y32 is associated with cancer, we propose that Y32 can be targeted by means of small therapeutics that can displace it from the nucleotide binding site, thus introducing intrinsic GTPase activity to RAS mutants, which cannot bind to GAP. Therefore, mimicking properties of H-RASG12P in RAS-centered drug discovery studies has the potential of improving success rates since it acts as a molecular switch per se.

Utilization of Biased G Protein-Coupled ReceptorSignaling towards Development of Safer andPersonalized Therapeutics.

G protein-coupled receptors (GPCRs) are involved in a wide variety of physiologicalprocesses. Therefore, approximately 40% of currently prescribed drugs have targeted this receptorfamily. Discovery of ß-arrestin mediated signaling and also separability of G protein and b-arrestinsignaling pathways have switched the research focus in the GPCR field towards development ofbiased ligands, which provide engagement of the receptor with a certain effector, thus enrichinga specific signaling pathway. In this review, we summarize possible factors that impact signalingprofiles of GPCRs such as oligomerization, drug treatment, disease conditions, genetic background,etc. along with relevant molecules that can be used to modulate signaling properties of GPCRssuch as allosteric or bitopic ligands, ions, aptamers and pepducins. Moreover, we also discuss theimportance of inclusion of pharmacogenomics and molecular dynamics simulations to achieve aholistic understanding of the relation between genetic background and structure and function ofGPCRs and GPCR-related proteins. Consequently, specific downstream signaling pathways can beenriched while those that bring unwanted side effects can be prevented on a patient-specific basis.This will improve studies that centered on development of safer and personalized therapeutics,thus alleviating the burden on economy and public health.

Computational Approaches in Antibody-drug Conjugate Optimization for Targeted Cancer Therapy.

Cancer has become one of the main leading causes of morbidity and mortality worldwide. One of the critical drawbacks of current cancer therapeutics has been the lack of the target-selectivity, as these drugs should have an effect exclusively on cancer cells while not perturbing healthy ones. In addition, their mechanism of action should be sufficiently fast to avoid the invasion of neighbouring healthy tissues by cancer cells. The use of conventional chemotherapeutic agents and other traditional therapies, such as surgery and radiotherapy, leads to off-target interactions with serious side effects. In this respect, recently developed target-selective Antibody-Drug Conjugates (ADCs) are more effective than traditional therapies, presumably due to their modular structures that combine many chemical properties simultaneously. In particular, ADCs are made up of three different units: a highly selective Monoclonal antibody (Mab) which is developed against a tumour-associated antigen, the payload (cytotoxic agent), and the linker. The latter should be stable in circulation while allowing the release of the cytotoxic agent in target cells. The modular nature of these drugs provides a platform to manipulate and improve selectivity and the toxicity of these molecules independently from each other. This in turn leads to generation of second- and third-generation ADCs, which have been more effective than the previous ones in terms of either selectivity or toxicity or both. Development of ADCs with improved efficacy requires knowledge at the atomic level regarding the structure and dynamics of the molecule. As such, we reviewed all the most recent computational methods used to attain all-atom description of the structure, energetics and dynamics of these systems. In particular, this includes homology modelling, molecular docking and refinement, atomistic and coarse-grained molecular dynamics simulations, principal component and cross-correlation analysis. The full characterization of the structure-activity relationship devoted to ADCs is critical for antibody-drug conjugate research and development.

Prediction and Targeting of Interaction Interfaces in G-protein Coupled Receptor Oligomers.

BACKGROUND: Communication within a protein complex is mediated by physical interactions made among the protomers. Evidence for both the allosteric regulation present among the protomers of the protein oligomer and of the direct effect of membrane composition on this regulation has made it essential to investigate the underlying molecular mechanism that drives oligomerization, the type of interactions present within the complex, and to determine the identity of the interaction interface. This knowledge allows a holistic understanding of dynamics and also modulation of the function of the resulting oligomers/signalling complexes. G-Protein-Coupled Receptors (GPCRs), which are targeted by 40% of currently prescribed drugs in the market, are widely involved in the formation of such physiological oligomers/signalling complexes. SCOPE: This review highlights the importance of studying Protein-Protein Interactions (PPI) by using a combination of data obtained from cutting-edge experimental and computational methods that were developed for this purpose. In particular, we focused on interaction interfaces found at GPCR oligomers as well as signalling complexes, since any problem associated with these interactions causes the onset of various crucial diseases. CONCLUSION: In order to have a holistic mechanistic understanding of allosteric PPIs that drive the formation of GPCR oligomers and also to determine the composition of interaction interfaces with respect to different membrane compositions, it is essential to combine both relevant experimental and computational data. In this way, efficient and specific targeting of these interaction interfaces in oligomers/ complexes can be achieved. Thus, effective therapeutic molecules with fewer side effects can be designed to modulate the function of these physiologically important receptor family.

Computational studies of G protein-coupled receptor complexes: Structure and dynamics.

G protein-coupled receptors (GPCRs) are ubiquitously expressed transmembrane proteins associated with a wide range of diseases such as Alzheimer's, Parkinson, schizophrenia, and also implicated in in several abnormal heart conditions. As such, this family of receptors is regarded as excellent drug targets. However, due to the high number of intracellular signaling partners, these receptors have a complex interaction networks and it becomes challenging to modulate their function. Experimentally determined structures give detailed information on the salient structural properties of these signaling complexes but they are far away from providing mechanistic insights into the underlying process. This chapter presents some of the computational tools, namely molecular dynamics, molecular docking, and molecular modeling and related analyses methods that have been used to complement experimental findings.