scholarly journals Structure-based inhibitor design of mutant RAS proteins—a paradigm shift

2020 ◽  
Vol 39 (4) ◽  
pp. 1091-1105
Author(s):  
Kinga Nyíri ◽  
Gergely Koppány ◽  
Beáta G. Vértessy
Keyword(s):  
Covalent Modification ◽  
Small Gtpase ◽  
Drug Binding ◽  
Ras Proteins ◽  
Structure Based Drug Design ◽  
Lung Cancers ◽  
Long Time ◽  
Binding Pockets ◽  
Cancer Types ◽  
Kras Protein

AbstractAs a member of small GTPase family, KRAS protein is a key physiological modulator of various cellular activities including proliferation. However, mutations of KRAS present in numerous cancer types, most frequently in pancreatic (> 60%), colorectal (> 40%), and lung cancers, drive oncogenic processes through overactivation of proliferation. The G12C mutation of KRAS protein is especially abundant in the case of these types of malignancies. Despite its key importance in human disease, KRAS was assumed to be non-druggable for a long time since the protein seemingly lacks potential drug-binding pockets except the nucleotide-binding site, which is difficult to be targeted due to the high affinity of KRAS for both GDP and GTP. Recently, a new approach broke the ice and provided evidence that upon covalent targeting of the G12C mutant KRAS, a highly dynamic pocket was revealed. This novel targeting is especially important since it serves with an inherent solution for drug selectivity. Based on these results, various structure-based drug design projects have been launched to develop selective KRAS mutant inhibitors. In addition to the covalent modification strategy mostly applicable for G12C mutation, different innovative solutions have been suggested for the other frequently occurring oncogenic G12 mutants. Here we summarize the latest advances of this field, provide perspectives for novel approaches, and highlight the special properties of KRAS, which might issue some new challenges.

Use of Molecular Dynamics Simulations in Structure-Based Drug Discovery

2019 ◽  
Vol 25 (31) ◽  
pp. 3339-3349 ◽  
Author(s):  
Indrani Bera ◽  
Pavan V. Payghan
Keyword(s):  
Molecular Dynamics ◽  
Drug Discovery ◽  
Molecular Dynamics Simulations ◽  
Drug Target ◽  
Structural Information ◽  
Drug Binding ◽  
Structure Based Drug Design ◽  
Drug Designing ◽  
Target Binding ◽  
Dynamics Simulations

Background: Traditional drug discovery is a lengthy process which involves a huge amount of resources. Modern-day drug discovers various multidisciplinary approaches amongst which, computational ligand and structure-based drug designing methods contribute significantly. Structure-based drug designing techniques require the knowledge of structural information of drug target and drug-target complexes. Proper understanding of drug-target binding requires the flexibility of both ligand and receptor to be incorporated. Molecular docking refers to the static picture of the drug-target complex(es). Molecular dynamics, on the other hand, introduces flexibility to understand the drug binding process. Objective: The aim of the present study is to provide a systematic review on the usage of molecular dynamics simulations to aid the process of structure-based drug design. Method: This review discussed findings from various research articles and review papers on the use of molecular dynamics in drug discovery. All efforts highlight the practical grounds for which molecular dynamics simulations are used in drug designing program. In summary, various aspects of the use of molecular dynamics simulations that underline the basis of studying drug-target complexes were thoroughly explained. Results: This review is the result of reviewing more than a hundred papers. It summarizes various problems that use molecular dynamics simulations. Conclusion: The findings of this review highlight how molecular dynamics simulations have been successfully implemented to study the structure-function details of specific drug-target complexes. It also identifies the key areas such as stability of drug-target complexes, ligand binding kinetics and identification of allosteric sites which have been elucidated using molecular dynamics simulations.

scholarly journals Unveiling the Dynamics of KRAS4b on Lipid Model Membranes

2021 ◽  
Author(s):  
Cesar A. López ◽  
Animesh Agarwal ◽  
Que N. Van ◽  
Andrew G. Stephen ◽  
S. Gnanakaran
Keyword(s):  
Molecular Mechanisms ◽  
Hypervariable Region ◽  
Small Gtpase ◽  
Model Systems ◽  
Coarse Grained ◽  
Regulatory Function ◽  
Limited Information ◽  
Long Time ◽  
Cell Growth And Differentiation ◽  
State Models

AbstractSmall GTPase proteins are ubiquitous and responsible for regulating several processes related to cell growth and differentiation. Mutations that stabilize their active state can lead to uncontrolled cell proliferation and cancer. Although these proteins are well characterized at the cellular scale, the molecular mechanisms governing their functions are still poorly understood. In addition, there is limited information about the regulatory function of the cell membrane which supports their activity. Thus, we have studied the dynamics and conformations of the farnesylated KRAS4b in various membrane model systems, ranging from binary fluid mixtures to heterogeneous raft mimics. Our approach combines long time-scale coarse-grained (CG) simulations and Markov state models to dissect the membrane-supported dynamics of KRAS4b. Our simulations reveal that protein dynamics is mainly modulated by the presence of anionic lipids and to some extent by the nucleotide state (activation) of the protein. In addition, our results suggest that both the farnesyl and the polybasic hypervariable region (HVR) are responsible for its preferential partitioning within the liquid-disordered (Ld) domains in membranes, potentially enhancing the formation of membrane-driven signaling platforms. Graphic Abstract

scholarly journals Breaking the next Cryo-EM resolution barrier – Atomic resolution determination of proteins!

2020 ◽  
Author(s):  
Ka Man Yip ◽  
Niels Fischer ◽  
Elham Paknia ◽  
Ashwin Chari ◽  
Holger Stark
Keyword(s):  
Protein Function ◽  
Model Building ◽  
Technological Development ◽  
Drug Binding ◽  
Single Atom ◽  
Biological Macromolecules ◽  
Direct Visualization ◽  
Hydrogen Atoms ◽  
Structure Based Drug Design ◽  
Electron Microscopes

SummarySingle particle cryo-EM is a powerful method to solve the three-dimensional structures of biological macromolecules. The technological development of electron microscopes, detectors, automated procedures in combination with user friendly image processing software and ever-increasing computational power have made cryo-EM a successful and largely expanding technology over the last decade. At resolutions better than 4 Å, atomic model building starts becoming possible but the direct visualization of true atomic positions in protein structure determination requires significantly higher (< 1.5 Å) resolution, which so far could not be attained by cryo-EM. The direct visualization of atom positions is essential for understanding protein-catalyzed chemical reaction mechanisms and to study drug-binding and -interference with protein function. Here we report a 1.25 Å resolution structure of apoferritin obtained by cryo-EM with a newly developed electron microscope providing unprecedented structural details. Our apoferritin structure has almost twice the 3D information content of the current world record reconstruction (at 1.54 Å resolution 1). For the first time in cryo-EM we can visualize individual atoms in a protein, see density for hydrogen atoms and single atom chemical modifications. Beyond the nominal improvement in resolution we can also show a significant improvement in quality of the cryo-EM density map which is highly relevant for using cryo-EM in structure-based drug design.

scholarly journals Automated Classification of Lung Cancer Types from Cytological Images Using Deep Convolutional Neural Networks

2017 ◽  
Vol 2017 ◽  
pp. 1-6 ◽  
Author(s):  
Atsushi Teramoto ◽  
Tetsuya Tsukamoto ◽  
Yuka Kiriyama ◽  
Hiroshi Fujita
Keyword(s):  
Lung Cancer ◽  
Cell Carcinoma ◽  
Graphics Processing Unit ◽  
Processing Unit ◽  
Automated Classification ◽  
Deep Convolutional Neural Networks ◽  
Lung Cancers ◽  
Microscopic Images ◽  
Cancer Types

Lung cancer is a leading cause of death worldwide. Currently, in differential diagnosis of lung cancer, accurate classification of cancer types (adenocarcinoma, squamous cell carcinoma, and small cell carcinoma) is required. However, improving the accuracy and stability of diagnosis is challenging. In this study, we developed an automated classification scheme for lung cancers presented in microscopic images using a deep convolutional neural network (DCNN), which is a major deep learning technique. The DCNN used for classification consists of three convolutional layers, three pooling layers, and two fully connected layers. In evaluation experiments conducted, the DCNN was trained using our original database with a graphics processing unit. Microscopic images were first cropped and resampled to obtain images with resolution of 256 × 256 pixels and, to prevent overfitting, collected images were augmented via rotation, flipping, and filtering. The probabilities of three types of cancers were estimated using the developed scheme and its classification accuracy was evaluated using threefold cross validation. In the results obtained, approximately 71% of the images were classified correctly, which is on par with the accuracy of cytotechnologists and pathologists. Thus, the developed scheme is useful for classification of lung cancers from microscopic images.

Fluorescent nanoparticle sensors with tailor-made recognition units and proximate fluorescent reporter groups

2018 ◽  
Vol 42 (12) ◽  
pp. 9377-9380 ◽  
Author(s):  
Xiaoyu Xing ◽  
Yan Zhao
Keyword(s):  
Molecular Imprinting ◽  
Binding Pocket ◽  
Fluorescent Sensors ◽  
Covalent Modification ◽  
Fluorescent Reporter ◽  
Binding Pockets ◽  
Fluorescent Nanoparticle

Molecular imprinting in micelles followed by covalent modification of the binding pocket yielded fluorescent sensors with precisely constructed binding pockets.

scholarly journals Interplay Between Mitochondrial Peroxiredoxins and ROS in Cancer Development and Progression

2019 ◽  
Vol 20 (18) ◽  
pp. 4407 ◽  
Author(s):  
Tayaba Ismail ◽  
Youni Kim ◽  
Hongchan Lee ◽  
Dong-Seok Lee ◽  
Hyun-Shik Lee
Keyword(s):  
Redox Balance ◽  
Cancer Development ◽  
Cancer Therapies ◽  
Lung Cancers ◽  
Micro Rnas ◽  
Cancer Types ◽  
High Level ◽  
Cancerous Cells ◽  
Ros Scavengers ◽  
Cellular Organelles

Mitochondria are multifunctional cellular organelles that are major producers of reactive oxygen species (ROS) in eukaryotes; to maintain the redox balance, they are supplemented with different ROS scavengers, including mitochondrial peroxiredoxins (Prdxs). Mitochondrial Prdxs have physiological and pathological significance and are associated with the initiation and progression of various cancer types. In this review, we have focused on signaling involving ROS and mitochondrial Prdxs that is associated with cancer development and progression. An upregulated expression of Prdx3 and Prdx5 has been reported in different cancer types, such as breast, ovarian, endometrial, and lung cancers, as well as in Hodgkin’s lymphoma and hepatocellular carcinoma. The expression of Prdx3 and Prdx5 in different types of malignancies involves their association with different factors, such as transcription factors, micro RNAs, tumor suppressors, response elements, and oncogenic genes. The microenvironment of mitochondrial Prdxs plays an important role in cancer development, as cancerous cells are equipped with a high level of antioxidants to overcome excessive ROS production. However, an increased production of Prdx3 and Prdx5 is associated with the development of chemoresistance in certain types of cancers and it leads to further complications in cancer treatment. Understanding the interplay between mitochondrial Prdxs and ROS in carcinogenesis can be useful in the development of anticancer drugs with better proficiency and decreased resistance. However, more targeted studies are required for exploring the tumor microenvironment in association with mitochondrial Prdxs to improve the existing cancer therapies and drug development.

scholarly journals Drugging an undruggable pocket on KRAS

2019 ◽  
Vol 116 (32) ◽  
pp. 15823-15829 ◽  
Author(s):  
Dirk Kessler ◽  
Michael Gmachl ◽  
Andreas Mantoulidis ◽  
Laetitia J. Martin ◽  
Andreas Zoephel ◽  
...  
Keyword(s):  
Molecular Switches ◽  
Small Gtpases ◽  
Ras Proteins ◽  
Structure Based Drug Design ◽  
Downstream Signaling ◽  
Activating Mutations ◽  
Ras Genes ◽  
Mutant Cells ◽  
Drug Discovery Target ◽  
Micromolar Range

The 3 human RAS genes, KRAS, NRAS, and HRAS, encode 4 different RAS proteins which belong to the protein family of small GTPases that function as binary molecular switches involved in cell signaling. Activating mutations in RAS are among the most common oncogenic drivers in human cancers, with KRAS being the most frequently mutated oncogene. Although KRAS is an excellent drug discovery target for many cancers, and despite decades of research, no therapeutic agent directly targeting RAS has been clinically approved. Using structure-based drug design, we have discovered BI-2852 (1), a KRAS inhibitor that binds with nanomolar affinity to a pocket, thus far perceived to be “undruggable,” between switch I and II on RAS; 1 is mechanistically distinct from covalent KRASG12C inhibitors because it binds to a different pocket present in both the active and inactive forms of KRAS. In doing so, it blocks all GEF, GAP, and effector interactions with KRAS, leading to inhibition of downstream signaling and an antiproliferative effect in the low micromolar range in KRAS mutant cells. These findings clearly demonstrate that this so-called switch I/II pocket is indeed druggable and provide the scientific community with a chemical probe that simultaneously targets the active and inactive forms of KRAS.

scholarly journals Analysis of RAS protein interactions in living cells reveals a mechanism for pan-RAS depletion by membrane-targeted RAS binders

2020 ◽  
Vol 117 (22) ◽  
pp. 12121-12130
Author(s):  
Yao-Cheng Li ◽  
Nikki K. Lytle ◽  
Seth T. Gammon ◽  
Luke Wang ◽  
Tikvah K. Hayes ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Developmental Disorders ◽  
Degradation Pathway ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Ras Proteins ◽  
Ras Protein ◽  
Protein Levels ◽  
Ras Family ◽  
Ras Isoforms

HRAS, NRAS, and KRAS4A/KRAS4B comprise the RAS family of small GTPases that regulate signaling pathways controlling cell proliferation, differentiation, and survival. RAS pathway abnormalities cause developmental disorders and cancers. We found that KRAS4B colocalizes on the cell membrane with other RAS isoforms and a subset of prenylated small GTPase family members using a live-cell quantitative split luciferase complementation assay. RAS protein coclustering is mainly mediated by membrane association-facilitated interactions (MAFIs). Using the RAS–RBD (CRAF RAS binding domain) interaction as a model system, we showed that MAFI alone is not sufficient to induce RBD-mediated RAS inhibition. Surprisingly, we discovered that high-affinity membrane-targeted RAS binding proteins inhibit RAS activity and deplete RAS proteins through an autophagosome–lysosome-mediated degradation pathway. Our results provide a mechanism for regulating RAS activity and protein levels, a more detailed understanding of which should lead to therapeutic strategies for inhibiting and depleting oncogenic RAS proteins.

scholarly journals Multidrug Resistance in Neisseria gonorrhoeae: Identification of Functionally Important Residues in the MtrD Efflux Protein

mBio ◽  
2019 ◽  
Vol 10 (6) ◽  
Author(s):  
Mohsen Chitsaz ◽  
Lauren Booth ◽  
Mitchell T. Blyth ◽  
Megan L. O’Mara ◽  
Melissa H. Brown
Keyword(s):  
Molecular Dynamics ◽  
Multidrug Resistance ◽  
Neisseria Gonorrhoeae ◽  
Molecular Dynamics Simulations ◽  
Docking Studies ◽  
Drug Binding ◽  
Multidrug Efflux ◽  
Content Type ◽  
Binding Pockets ◽  
Dynamics Simulations

ABSTRACT A key mechanism that Neisseria gonorrhoeae uses to achieve multidrug resistance is the expulsion of structurally different antimicrobials by the MtrD multidrug efflux protein. MtrD resembles the homologous Escherichia coli AcrB efflux protein with several common structural features, including an open cleft containing putative access and deep binding pockets proposed to interact with substrates. A highly discriminating N. gonorrhoeae strain, with the MtrD and NorM multidrug efflux pumps inactivated, was constructed and used to confirm and extend the substrate profile of MtrD to include 14 new compounds. The structural basis of substrate interactions with MtrD was interrogated by a combination of long-timescale molecular dynamics simulations and docking studies together with site-directed mutagenesis of selected residues. Of the MtrD mutants generated, only one (S611A) retained a wild-type (WT) resistance profile, while others (F136A, F176A, I605A, F610A, F612C, and F623C) showed reduced resistance to different antimicrobial compounds. Docking studies of eight MtrD substrates confirmed that many of the mutated residues play important nonspecific roles in binding to these substrates. Long-timescale molecular dynamics simulations of MtrD with its substrate progesterone showed the spontaneous binding of the substrate to the access pocket of the binding cleft and its subsequent penetration into the deep binding pocket, allowing the permeation pathway for a substrate through this important resistance mechanism to be identified. These findings provide a detailed picture of the interaction of MtrD with substrates that can be used as a basis for rational antibiotic and inhibitor design. IMPORTANCE With over 78 million new infections globally each year, gonorrhea remains a frustratingly common infection. Continuous development and spread of antimicrobial-resistant strains of Neisseria gonorrhoeae, the causative agent of gonorrhea, have posed a serious threat to public health. One of the mechanisms in N. gonorrhoeae involved in resistance to multiple drugs is performed by the MtrD multidrug resistance efflux pump. This study demonstrated that the MtrD pump has a broader substrate specificity than previously proposed and identified a cluster of residues important for drug binding and translocation. Additionally, a permeation pathway for the MtrD substrate progesterone actively moving through the protein was determined, revealing key interactions within the putative MtrD drug binding pockets. Identification of functionally important residues and substrate-protein interactions of the MtrD protein is crucial to develop future strategies for the treatment of multidrug-resistant gonorrhea.

Chemical parameters influencing fine-tuning in the binding of macrolide antibiotics to the ribosomal tunnel

2007 ◽  
Vol 79 (6) ◽  
pp. 955-968 ◽  
Author(s):  
Erez Pyetan ◽  
David Baram ◽  
Tamar Auerbach-Nevo ◽  
Ada Yonath
Keyword(s):  
Deinococcus Radiodurans ◽  
Bacterial Species ◽  
Ribosomal Subunit ◽  
Binding Pocket ◽  
Fine Tuning ◽  
23S Rrna ◽  
Structure Based Drug Design ◽  
Altered Level ◽  
Ribosomal Tunnel ◽  
Binding Pockets

In comparison to existing structural, biochemical, and therapeutical data, the crystal structures of large ribosomal subunit from the eubacterial pathogen model Deinococcus radiodurans in complex with the 14-membered macrolides erythromycylamine, RU69874, and the 16-membered macrolide josamycin, highlighted the similarities and differences in macrolides binding to the ribosomal tunnel. The three compounds occupy the macrolide binding pocket with their desosamine or mycaminose aminosugar, the C4-C7 edge of the macrolactone ring and the cladinose sugar sharing similar positions and orientations, although the latter, known to be unnecessary for antibiotic activity, displays fewer contacts. The macrolactone ring displays altogether few contacts with the ribosome and can, therefore, tilt in order to optimize its interaction with the 23S rRNA. In addition to their contacts with nucleotides of domain V of the 23S RNA, erythromycylamine and RU69874 interact with domain II nucleotide U790, and RU69874 also reaches van der Waals distance from A752, in a fashion similar to that observed for the ketolides telithromycin and cethromycin. The variability in the sequences and consequently the diversity of the conformations of macrolide binding pockets in various bacterial species can explain the drug's altered level of effectiveness on different organisms and is thus an important factor in structure-based drug design.

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