Quinoline-based Protein–protein Interaction Inhibitors of LEDGF/p75 and HIV Integrase: An In Silico Study

2019 ◽  
Vol 18 (32) ◽  
pp. 2800-2815 ◽  
Author(s):  
Nisha Chhokar ◽  
Sourav Kalra ◽  
Monika Chauhan ◽  
Anjana Munshi ◽  
Raj Kumar
Keyword(s):  
Protein Interactions ◽  
Biological Data ◽  
Strand Transfer ◽  
Hiv Integrase ◽  
Protein Protein Interactions ◽  
Allosteric Site ◽  
Protein Protein Interaction ◽  
In Silico Study ◽  
Cellular Cofactor

The failure of the Integrase Strand Transfer Inhibitors (INSTIs) due to the mutations occurring at the catalytic site of HIV integrase (IN) has led to the design of allosteric integrase inhibitors (ALLINIs). Lens epithelium derived growth factor (LEDGF/p75) is the host cellular cofactor which helps chaining IN to the chromatin. The protein-protein interactions (PPIs) were observed at the allosteric site (LEDGF/p75 binding domain) between LEDGF/p75 of the host cell and IN of virus. In recent years, many small molecules such as CX04328, CHIBA-3053 and CHI-104 have been reported as LEDGF/p75-IN interaction inhibitors (LEDGINs). LEDGINs have emerged as promising therapeutics to halt the PPIs by binding at the interface of both the proteins. In the present work, we correlated the docking scores for the reported LEDGINs containing quinoline scaffold with the in vitro biological data. The hierarchal clustering method was used to divide the compounds into test and training set. The robustness of the generated model was validated by q2 and r2 for the predicted set of compounds. The generated model between the docking score and biological data was assessed to predict the activity of the hits (quinoline scaffold) obtained from virtual screening of LEDGINs providing their structureactivity relationships to aim for the generation of potent agents.

scholarly journals Evolutionary diversification of protein–protein interactions by interface add-ons

2017 ◽  
Vol 114 (40) ◽  
pp. E8333-E8342 ◽  
Author(s):  
Maximilian G. Plach ◽  
Florian Semmelmann ◽  
Florian Busch ◽  
Markus Busch ◽  
Leonhard Heizinger ◽  
...  
Keyword(s):  
Protein Interaction ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Evolutionary Strategy ◽  
Structural Level ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Evolutionary Diversification

Cells contain a multitude of protein complexes whose subunits interact with high specificity. However, the number of different protein folds and interface geometries found in nature is limited. This raises the question of how protein–protein interaction specificity is achieved on the structural level and how the formation of nonphysiological complexes is avoided. Here, we describe structural elements called interface add-ons that fulfill this function and elucidate their role for the diversification of protein–protein interactions during evolution. We identified interface add-ons in 10% of a representative set of bacterial, heteromeric protein complexes. The importance of interface add-ons for protein–protein interaction specificity is demonstrated by an exemplary experimental characterization of over 30 cognate and hybrid glutamine amidotransferase complexes in combination with comprehensive genetic profiling and protein design. Moreover, growth experiments showed that the lack of interface add-ons can lead to physiologically harmful cross-talk between essential biosynthetic pathways. In sum, our complementary in silico, in vitro, and in vivo analysis argues that interface add-ons are a practical and widespread evolutionary strategy to prevent the formation of nonphysiological complexes by specializing protein–protein interactions.

scholarly journals Screening for Antiviral Inhibitors of the HIV Integrase—LEDGF/p75 Interaction Using the AlphaScreen™ Luminescent Proximity Assay

2008 ◽  
Vol 13 (5) ◽  
pp. 406-414 ◽  
Author(s):  
Yan Hou ◽  
Debra E. Mcguinness ◽  
Andrew J. Prongay ◽  
Boris Feld ◽  
Paul Ingravallo ◽  
...  
Keyword(s):  
Amino Acid ◽  
Strand Transfer ◽  
Hiv Integrase ◽  
Catalytic Core ◽  
Core Domain ◽  
Protein Protein Interaction ◽  
New Class ◽  
Cellular Cofactor ◽  
Hiv Integrase Inhibitors ◽  
Dna Complex

Small-molecule inhibitors of HIV integrase (HIV IN) have emerged as a promising new class of antivirals for the treatment of HIV/AIDS. The compounds currently approved or in clinical development specifically target HIV DNA integration and were identified using strand-transfer assays targeting the HIV IN/viral DNA complex. The authors have developed a second biochemical assay for identification of HIV integrase inhibitors, targeting the interaction between HIV IN and the cellular cofactor LEDGF/p75. They developed a luminescent proximity assay (AlphaScreen™) designed to measure the association of the 80-amino-acid integrase binding domain of LEDGF/p75 with the 163-amino-acid catalytic core domain of HIV IN. This assay proved to be quite robust (with a Z′ factor of 0.84 in screening libraries arrayed as orthogonal mixtures) and successfully identified several compounds specific for this protein-protein interaction. ( Journal of Biomolecular Screening 2008:406-414)

scholarly journals Mapping of the Hepatitis B Virus Reverse Transcriptase TP and RT Domains by Transcomplementation for Nucleotide Priming and by Protein-Protein Interaction

1999 ◽  
Vol 73 (3) ◽  
pp. 1885-1893 ◽  
Author(s):  
Robert E. Lanford ◽  
Young-Ho Kim ◽  
Helen Lee ◽  
Lena Notvall ◽  
Burton Beames
Keyword(s):  
Amino Acids ◽  
Reverse Transcriptase ◽  
Protein Interaction ◽  
Protein Interactions ◽  
Rnase H ◽  
Genome Replication ◽  
Protein Protein Interactions ◽  
Stem Loop ◽  
Protein Protein Interaction

ABSTRACT Hepadnavirus polymerases initiate reverse transcription in a protein-primed reaction. We previously described a complementation assay for analysis of the roles of the TP and RT domains of HBV reverse transcriptase (pol) in the priming reaction. Independently expressed TP and RT domains form a complex functional for in vitro priming reactions. To map the minimal functional TP and RT domains, we prepared baculoviruses expressing amino- and carboxyl-terminal deletions of both the TP and RT domains and analyzed the proteins for the ability to participate in transcomplementation for the priming reaction. The minimal TP domain spanned amino acids 20 to 175; however, very little activity was observed without a TP domain spanning amino acids 1 to 199. The minimal RT domain spanned amino acids 300 to 775; however, little activity was observed unless the carboxyl end of the RT domain extended to amino acid 800. Thus, most of the RNase H domain was required. In previous studies, we observed a TP inhibitory domain between amino acids 199 and 344. The current analysis narrowed this domain to residues 300 to 334, which is a portion of the minimal RT domain. In addition, the ability of TP and RT deletion mutants to form stable TP-RT complexes was examined in coimmunoprecipitation assays. The minimal TP and RT domains capable of protein-protein interaction were considerably smaller than the domains required for functional interaction in the transcomplementation assays, and unlike priming activity, TP-RT interaction did not require the epsilon RNA stem-loop. These studies help to further define the complex protein-protein interactions required in HBV genome replication.

scholarly journals BRET-based RAS biosensors that show a novel small molecule is an inhibitor of RAS-effector protein-protein interactions

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Nicolas Bery ◽  
Abimael Cruz-Migoni ◽  
Carole JR Bataille ◽  
Camilo E Quevedo ◽  
Hanna Tulmin ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Human Cancers ◽  
Ras Family ◽  
Ras Inhibitors ◽  
In Cells

The RAS family of proteins is amongst the most highly mutated in human cancers and has so far eluded drug therapy. Currently, much effort is being made to discover mutant RAS inhibitors and in vitro screening for RAS-binding drugs must be followed by cell-based assays. Here, we have developed a robust set of bioluminescence resonance energy transfer (BRET)-based RAS biosensors that enable monitoring of RAS-effector interaction inhibition in living cells. These include KRAS, HRAS and NRAS and a variety of different mutations that mirror those found in human cancers with the major RAS effectors such as CRAF, PI3K and RALGDS. We highlighted the utility of these RAS biosensors by showing a RAS-binding compound is a potent pan-RAS-effector interactions inhibitor in cells. The RAS biosensors represent a useful tool to investigate and characterize the potency of anti-RAS inhibitors in cells and more generally any RAS protein-protein interaction (PPI) in cells.

scholarly journals Prediction of Protein-Protein Interactions Related to Protein Complexes Based on Protein Interaction Networks

2015 ◽  
Vol 2015 ◽  
pp. 1-9
Author(s):  
Peng Liu ◽  
Lei Yang ◽  
Daming Shi ◽  
Xianglong Tang
Keyword(s):  
Protein Interaction ◽  
Protein Interactions ◽  
Protein Complexes ◽  
Statistical Tests ◽  
Interaction Network ◽  
Biological Data ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Small Parts

A method for predicting protein-protein interactions based on detected protein complexes is proposed to repair deficient interactions derived from high-throughput biological experiments. Protein complexes are pruned and decomposed into small parts based on the adaptivek-cores method to predict protein-protein interactions associated with the complexes. The proposed method is adaptive to protein complexes with different structure, number, and size of nodes in a protein-protein interaction network. Based on different complex sets detected by various algorithms, we can obtain different prediction sets of protein-protein interactions. The reliability of the predicted interaction sets is proved by using estimations with statistical tests and direct confirmation of the biological data. In comparison with the approaches which predict the interactions based on the cliques, the overlap of the predictions is small. Similarly, the overlaps among the predicted sets of interactions derived from various complex sets are also small. Thus, every predicted set of interactions may complement and improve the quality of the original network data. Meanwhile, the predictions from the proposed method replenish protein-protein interactions associated with protein complexes using only the network topology.

Disruption of protein–protein interactions: hot spot detection, structure-based virtual screening and in vitro testing for the anti-cancer drug target – survivin

RSC Advances ◽  
2016 ◽  
Vol 6 (38) ◽  
pp. 31947-31959 ◽  
Author(s):  
Sailu Sarvagalla ◽  
Chun Hei Antonio Cheung ◽  
Ju-Ya Tsai ◽  
Hsing Pang Hsieh ◽  
Mohane Selvaraj Coumar
Keyword(s):  
Protein Interactions ◽  
Drug Target ◽  
Hot Spot ◽  
Cancer Drug ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Spot Detection ◽  
Anti Cancer ◽  
Interaction Interface

Hot spot detection at the protein–protein interaction interface using computational tools helped to identify indinavir as survivin inhibitor.

scholarly journals Photoproximity Profiling of Protein-Protein Interactions in Cells

2019 ◽  
Author(s):  
David C. McCutcheon ◽  
Gihoon Lee ◽  
Anthony Carlos ◽  
Jeffrey E. Montgomery ◽  
Raymond E. Moellering
Keyword(s):  
Protein Interaction ◽  
Protein Interactions ◽  
Live Cells ◽  
High Temporal Resolution ◽  
Protein Protein Interactions ◽  
Redox Stress ◽  
Covalent Interaction ◽  
Protein Protein Interaction ◽  
In Cells

ABSTRACTWe report a novel photoproximity protein interaction (PhotoPPI) profiling method to map protein-protein interactions in vitro and in live cells. This approach utilizes a bioorthogonal, multifunctional chemical probe that can be targeted to a genetically encoded protein of interest (POI) through a modular SNAP-Tag/benzylguanine covalent interaction. A first generation photoproximity probe, PP1, responds to 365 nm light to simultaneously cleave a central nitroveratryl linker and a peripheral diazirine group, resulting in diffusion of a highly reactive carbene nucleophile away from the POI. We demonstrate facile probe loading, and subsequent interaction- and light-dependent proximal labeling of a model protein-protein interaction (PPI) in vitro. Integration of the PhotoPPI workflow with quantitative LC-MS/MS enabled un-biased interaction mapping for the redox regulated sensor protein, KEAP1, for the first time in live cells. We validated known and novel interactions between KEAP1 and the proteins PGAM5 and HK2, among others, under basal cellular conditions. By contrast, comparison of PhotoPPI profiles in cells experiencing metabolic or redox stress confirmed that KEAP1 sheds many basal interactions and becomes associated with known lysosomal trafficking and proteolytic proteins like SQSTM1, CTSD and LGMN. Together, these data establish PhotoPPI as a method capable of tracking the dynamic sub-cellular and protein interaction “social network” of a redox-sensitive protein in cells with high temporal resolution.SYNOPSIS TOC

Probing the 14-3-3 Isoform-Specificity Profile of Interactions Stabilized by Fusicoccin A

2020 ◽  
Author(s):  
James Frederich ◽  
Ananya Sengupta ◽  
Josue Liriano ◽  
Ewa A. Bienkiewicz ◽  
Brian G. Miller
Keyword(s):  
Protein Interactions ◽  
Neurological Diseases ◽  
Adapter Proteins ◽  
Protein Protein Interactions ◽  
Specific Expression ◽  
Individual Client ◽  
Isoform Specificity ◽  
Fusicoccin A

Fusicoccin A (FC) is a fungal phytotoxin that stabilizes protein–protein interactions (PPIs) between 14-3-3 adapter proteins and their phosphoprotein interaction partners. In recent years, FC has emerged as an important chemical probe of human 14-3-3 PPIs implicated in cancer and neurological diseases. These previous studies have established the structural requirements for FC-induced stabilization of 14-3-3·client phosphoprotein complexes; however, the effect of different 14-3-3 isoforms on FC activity has not been systematically explored. This is a relevant question for the continued development of FC variants because there are seven distinct isoforms of 14-3-3 in humans. Despite their remarkable sequence and structural similarities, a growing body of experimental evidence supports both tissue-specific expression of 14-3-3 isoforms and isoform-specific functions <i>in vivo</i>. Herein, we report the isoform-specificity profile of FC <i>in vitro</i>using recombinant human 14-3-3 isoforms and a focused library of fluorescein-labeled hexaphosphopeptides mimicking the C-terminal 14-3-3 recognition domains of client phosphoproteins targeted by FC in cell culture. Our results reveal modest isoform preferences for individual client phospholigands and demonstrate that FC differentially stabilizes PPIs involving 14-3-3s. Together, these data provide strong motivation for the development of non-natural FC variants with enhanced selectivity for individual 14-3-3 isoforms.

Discovery of New AKT1 Inhibitors by Combination of In silico Structure Based Virtual Screening Approaches and Biological Evaluations

2019 ◽  
Author(s):  
Filip Fratev ◽  
Denisse A. Gutierrez ◽  
Renato J. Aguilera ◽  
suman sirimulla
Keyword(s):  
Virtual Screening ◽  
Success Rate ◽  
Protein Interactions ◽  
In Silico ◽  
Biological Data ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
Screening Protocol ◽  
Screening Approaches

AKT1 is emerging as a useful target for treating cancer. Herein, we discovered a new set of ligands that inhibit the AKT1, as shown by in vitro binding and cell line studies, using a newly designed virtual screening protocol that combines structure-based pharmacophore and docking screens. Taking together with the biological data, the combination of structure based pharamcophore and docking methods demonstrated reasonable success rate in identifying new inhibitors (60-70%) proving the success of aforementioned approach. A detail analysis of the ligand-protein interactions was performed explaining observed activities.<br>

An efficient transient assays system using Agrobacterium-mediated transformation of onion (Allium cepa) epidermal cells

2020 ◽  
Vol 80 (03) ◽  
Author(s):  
Yu-Miao Zhang ◽  
Jun Wang ◽  
Tao Wu
Keyword(s):  
Subcellular Localization ◽  
Protein Interaction ◽  
Protein Interactions ◽  
Epidermal Cells ◽  
Cyclin Dependent Kinase ◽  
Protein Subcellular Localization ◽  
Protein Protein Interactions ◽  
Efficient System ◽  
Protein Protein Interaction ◽  
Onion Epidermal Cells

In this study, the Agrobacterium infection medium, infection duration, detergent, and cell density were optimized. The sorghum-based infection medium (SbIM), 10-20 min infection time, addition of 0.01% Silwet L-77, and Agrobacterium optical density at 600 nm (OD600), improved the competence of onion epidermal cells to support Agrobacterium infection at >90% efficiency. Cyclin-dependent kinase D-2 (CDKD-2) and cytochrome c-type biogenesis protein (CYCH), protein-protein interactions were localized. The optimized procedure is a quick and efficient system for examining protein subcellular localization and protein-protein interaction.

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