scholarly journals Structure of PDE3A–SLFN12 complex and structure-based design for a potent apoptosis inducer of tumor cells

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jie Chen ◽  
Nan Liu ◽  
Yinpin Huang ◽  
Yuanxun Wang ◽  
Yuxing Sun ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Catalytic Domain ◽  
Cultured Cells ◽  
Hydrophobic Interactions ◽  
Complex Structure ◽  
Protein A ◽  
Protein Translation ◽  
Protein Protein Interactions ◽  
Cryo Electron Microscopy ◽  
Apoptosis Inducer

AbstractMolecular glues are a class of small molecular drugs that mediate protein-protein interactions, that induce either the degradation or stabilization of target protein. A structurally diverse group of chemicals, including 17-β-estradiol (E2), anagrelide, nauclefine, and DNMDP, induces apoptosis by forming complexes with phosphodiesterase 3A (PDE3A) and Schlafen 12 protein (SLFN12). They do so by binding to the PDE3A enzymatic pocket that allows the compound-bound PDE3A to recruit and stabilize SLFN12, which in turn blocks protein translation, leading to apoptosis. In this work, we report the high-resolution cryo-electron microscopy structure of PDE3A-SLFN12 complexes isolated from cultured HeLa cells pre-treated with either anagrelide, or nauclefine, or DNMDP. The PDE3A-SLFN12 complexes exhibit a butterfly-like shape, forming a heterotetramer with these small molecules, which are packed in a shallow pocket in the catalytic domain of PDE3A. The resulting small molecule-modified interface binds to the short helix (E552-I558) of SLFN12 through hydrophobic interactions, thus “gluing” the two proteins together. Based on the complex structure, we designed and synthesized analogs of anagrelide, a known drug used for the treatment of thrombocytosis, to enhance their interactions with SLFN12, and achieved superior efficacy in inducing apoptosis in cultured cells as well as in tumor xenografts.

Complex Structure of PDE3A–SLFN12 and Structure-based Molecular Glue Design for Apoptosis of Tumor Cells

2021 ◽  
Author(s):  
Jie Chen ◽  
Nan Liu ◽  
Dianrong Li ◽  
Yuanxun Wang ◽  
Yuxing Sun ◽  
...  
Keyword(s):  
Cell Death ◽  
High Resolution ◽  
Tumor Cells ◽  
Protein Interactions ◽  
Catalytic Domain ◽  
Complex Structure ◽  
Protein Stabilization ◽  
Protein Protein Interactions ◽  
Essential Thrombocytosis ◽  
New Strategy

Abstract Molecular glue is a class of small molecular drugs that mediate the protein-protein interactions. It can induce target proteins degradation or stabilization. Anagrelide, as a known phosphodiesterase 3A (PDE3A) inhibitor, is a drug used for the treatment of essential thrombocytosis. Here, we report Anagrelide induces cell death directly binding to PDE3A, which stabilizes protein Schlafen 12 (SLFN12). The high-resolution cryo-EM maps indicates that the interaction of PDE3A and SLFN12 exhibits a butterfly-like shape, to form a heterotetramer. Structure analysis showed that Anagrelide, as a new molecular glue, packs in a shallow pocket of PDE3A in the catalytic domain and the resulting modified interface binds SLFN12 through the short helix (E552-I558) of SLFN12. Based on the structure, we designed and synthesized the Anagrelide analogues with hydrophobic substitutes at 7-position. The p-tolyl substitution (compound A6) had the best improvement of around 20 folds, which IC50 is around 0.3 nM. Our studies revealed a new strategy to identify and develop a new molecular glue for targeted protein stabilization.

scholarly journals Mechanism of NanR gene repression and allosteric induction of bacterial sialic acid metabolism

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Christopher R. Horne ◽  
Hariprasad Venugopal ◽  
Santosh Panjikar ◽  
David M. Wood ◽  
Amy Henrickson ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Dna Binding ◽  
Protein Interactions ◽  
Acid Metabolism ◽  
Environmental Changes ◽  
Dna Interaction ◽  
Protein Protein Interactions ◽  
Nutrient Source ◽  
Cryo Electron Microscopy ◽  
Close Proximity

AbstractBacteria respond to environmental changes by inducing transcription of some genes and repressing others. Sialic acids, which coat human cell surfaces, are a nutrient source for pathogenic and commensal bacteria. The Escherichia coli GntR-type transcriptional repressor, NanR, regulates sialic acid metabolism, but the mechanism is unclear. Here, we demonstrate that three NanR dimers bind a (GGTATA)3-repeat operator cooperatively and with high affinity. Single-particle cryo-electron microscopy structures reveal the DNA-binding domain is reorganized to engage DNA, while three dimers assemble in close proximity across the (GGTATA)3-repeat operator. Such an interaction allows cooperative protein-protein interactions between NanR dimers via their N-terminal extensions. The effector, N-acetylneuraminate, binds NanR and attenuates the NanR-DNA interaction. The crystal structure of NanR in complex with N-acetylneuraminate reveals a domain rearrangement upon N-acetylneuraminate binding to lock NanR in a conformation that weakens DNA binding. Our data provide a molecular basis for the regulation of bacterial sialic acid metabolism.

scholarly journals EEF1A2 interacts with HSP90AB1 to promote lung adenocarcinoma metastasis via enhancing TGF-β/SMAD signalling

2021 ◽  
Author(s):  
Liqing Jia ◽  
Xiaolu Ge ◽  
Chao Du ◽  
Linna Chen ◽  
Yanhong Zhou ◽  
...  
Keyword(s):  
Lung Adenocarcinoma ◽  
Protein Interactions ◽  
Elongation Factor ◽  
Protein Translation ◽  
Protein Protein Interactions ◽  
Promising Target ◽  
Tgfβ Receptor ◽  
Mesenchymal Transformation

Abstract Background Eukaryotic protein translation elongation factor 1α2 (EEF1A2) is an oncogene that promotes the progression of breast and pancreatic cancer. In this study, we aimed to elucidate the oncogenic function of EEF1A2 in the metastasis of lung adenocarcinoma (LUAD). Methods Immunohistochemistry and western blot were used to study EEF1A2 expression levels in LUAD tissues and cells, respectively. The role of EEF1A2 in LUAD progression were investigated in vitro and in vivo. We identified potential EEF1A2-binding proteins by liquid chromatography-electrospray mass spectrometry (LC-MS)/MS. Protein–protein interactions were determined by immunofluorescence and co-immunoprecipitation (Co-IP). Results In this study, we report that EEF1A2 mediates the epithelial–mesenchymal transformation (EMT), to promote the metastasis of LUAD cells in vitro and in vivo. Moreover, EEF1A2 interacts with HSP90AB1 to increase TGFβ Receptor (TβR)-I, and TβRII expression, followed by enhanced SMAD3 and pSMAD3 expression and nuclear localisation, which promotes the EMT of LUAD cells. Overexpression of EEF1A2 in cancer tissues is associated with poor prognosis and short survival of patients with LUAD. Conclusions These findings underscore the molecular functions of EEF1A2 in LUAD metastasis and indicate that EEF1A2 represents a promising target in the treatment of aggressive LUAD.

scholarly journals The Role of Posttranslational Modifications in DNA Repair

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Miaomiao Bai ◽  
Dongdong Ti ◽  
Qian Mei ◽  
Jiejie Liu ◽  
Xin Yan ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Protein Interactions ◽  
Posttranslational Modifications ◽  
Genome Stability ◽  
Protein Localization ◽  
Complex Structure ◽  
Protein Protein Interactions ◽  
Regulate Protein

The human body is a complex structure of cells, which are exposed to many types of stress. Cells must utilize various mechanisms to protect their DNA from damage caused by metabolic and external sources to maintain genomic integrity and homeostasis and to prevent the development of cancer. DNA damage inevitably occurs regardless of physiological or abnormal conditions. In response to DNA damage, signaling pathways are activated to repair the damaged DNA or to induce cell apoptosis. During the process, posttranslational modifications (PTMs) can be used to modulate enzymatic activities and regulate protein stability, protein localization, and protein-protein interactions. Thus, PTMs in DNA repair should be studied. In this review, we will focus on the current understanding of the phosphorylation, poly(ADP-ribosyl)ation, ubiquitination, SUMOylation, acetylation, and methylation of six typical PTMs and summarize PTMs of the key proteins in DNA repair, providing important insight into the role of PTMs in the maintenance of genome stability and contributing to reveal new and selective therapeutic approaches to target cancers.

scholarly journals An integrative approach unveils a distal encounter site for rPTPε and phospho-Src complex formation

2021 ◽  
Author(s):  
Nadendla EswarKumar ◽  
Cheng-Han Yang ◽  
Sunilkumar Tewary ◽  
Yi-Qi Yeh ◽  
Hsiao-Ching Yang ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Protein Complex ◽  
Tyrosine Phosphatase ◽  
Complex Structure ◽  
Protein Crystallography ◽  
Md Simulations ◽  
Integrative Approach ◽  
Single Mutation ◽  
Protein Protein Interactions ◽  
Transient Nature

AbstractProtein tyrosine phosphatase: phospho-protein complex structure determination, which requires to understand how specificity is achieved at the protein level remains a significant challenge for protein crystallography and cryoEM due to the transient nature of binding interactions. Using rPTPεD1 and phospho-SrcKD as a model system, we established an integrative workflow involving protein crystallography, SAXS and pTyr-tailored MD simulations to reveal the complex formed between rPTPεD1 and phospho-SrcKD, revealing transient protein–protein interactions distal to the active site. To support our finding, we determined the associate rate between rPTPεD1 and phospho-SrcKD and showed that a single mutation on rPTPεD1 disrupts this transient interaction, resulting in the reduction of association rate and activity. Our simulations suggest that rPTPεD1 employs a binding mechanism involving conformational change prior to the engagement of cSrcKD. This integrative approach is applicable to other PTP: phospho-protein complex determination and is a general approach for elucidating transient protein surface interactions.

scholarly journals A Computational Approach for Protein-Protein Interactions of Bacterial Surface Layer Protein with Human Erb3 and αIIB-β3 Receptors

2021 ◽  
Vol 12 (1) ◽  
pp. 420-430
Keyword(s):  
Surface Layer ◽  
Molecular Docking ◽  
Protein Interactions ◽  
Protein A ◽  
Computational Approach ◽  
Microbial Interactions ◽  
Protein Protein Interactions ◽  
Epithelial Surface ◽  
Surface Receptors ◽  
Surface Layer Protein

Host microbial interactions had significant factors in maintains homeostasis and immune-related activity. One such interaction made by Lactobacillus sp. with Surface layer proteins (Slps) had been studied through a computational approach. Erb3 and αIIB-β3, which are epithelial surface layer receptors, are subjected to interact with the Slp homology model. Both cell surface receptors were subjected to interact through computational docking, followed by molecular dynamics simulations through the coarse-grain method to explore the conformational stability. Through the implementation of the molecular docking for the surface layer protein A, we have shown the surface layer protein A, protein-protein interactions are higher in cellular receptors with epidermal growth factor receptor at an -34.45 ΔG and -51.19 ΔG through molecular docking with Erb3 and αIIB-β3. This study shows the unique interaction of Slp with the epithelial surface receptors like Erb3 and αIIB-β3, which are multipurpose applications in microbial-based drug therapeutics.

Water-holding capacity and characterization of protein interactions in processed cheese

1995 ◽  
Vol 62 (3) ◽  
pp. 479-489 ◽  
Author(s):  
Sylvie Marchesseau ◽  
Jean-Louis Cuq
Keyword(s):  
Protein Interactions ◽  
Hydrophobic Interactions ◽  
Visual Assessment ◽  
Protein Protein Interactions ◽  
Water Release ◽  
Water Binding ◽  
Processed Cheese ◽  
Protein Gels ◽  
Higher Temperature ◽  
Water Holding

SummaryIn order to predict the oozing susceptibility of heat-induced milk protein gels such as processed cheeses during storage, ultracentrifugation forces are required to accelerate the expulsion of water from the gel structure. Two predictive methods have been tested. Direct ultracentrifugation of processed cheese was used to study effects of centrifugal compression on the water-holding ability. The water release with optimal parameters (time, temperature and ultracentrifugation force) from a freshly manufactured processed cheese was correlated with visual assessment of oozing after 6 months storage. A second method was based on the susceptibility of gelled proteins to resist the dissociating action of solutions of chemical agents such as SDS, urea, EDTA and 2-mercaptoethanol. Most of the protein sedimented by ultracentrifugation in the presence of SDS represented that still in complexed form. This correlated with optimal water binding by the processed cheese. Response-surface methods, used to optimize the ultracentrifugation variables and the composition of the dissociation solutions, showed that the dissociation test was best with a dispersion of processed cheese in SDS solution (10 g/l) at a ratio of 1:6 (w/v) at 20°C, ultracentrifuged at 86000 g and 20°C for 25 min. Analysis of processed cheese cooked at 115°C and then dissociated in different solutions showed various protein–protein interactions in the gel network, whereas hydrophobic interactions were the most important stabilizers of the protein matrix of cheese cooked at higher temperature.

scholarly journals Thyroid stimulating autoantibody M22 mimics TSH binding to the TSH receptor leucine rich domain: a comparative structural study of protein–protein interactions

2009 ◽  
Vol 42 (5) ◽  
pp. 381-395 ◽  
Author(s):  
R Núñez Miguel ◽  
J Sanders ◽  
D Y Chirgadze ◽  
J Furmaniak ◽  
B Rees Smith
Keyword(s):  
Crystal Structure ◽  
Protein Interactions ◽  
Hydrophobic Interactions ◽  
Human Monoclonal Antibody ◽  
Tsh Receptor ◽  
Salt Bridges ◽  
Protein Protein Interactions ◽  
Rich Domain ◽  
Candidate Target ◽  
Comparative Model

The TSH receptor (TSHR) ligands M22 (a thyroid stimulating human monoclonal antibody) and TSH, bind to the concave surface of the leucine rich repeats domain (LRD) of the TSHR and here, we show that M22 mimics closely the binding of TSH. We compared interactions produced by M22 with the TSHR in the M22–TSHR crystal structure (2.55 Å resolution) and produced by TSH with the TSHR in a TSH–TSHR comparative model. The crystal structure of the TSHR and a comparative model of TSH based on the crystal structure of FSH were used as components to build the TSH–TSHR model. This model was built based on the FSH–FSH receptor structure (2.9 Å) and then the structure of the TSHR in the model was replaced by the TSHR crystal structure. The analysis shows that M22 light chain mimics the TSHβ chain in its interaction with TSHR LRD, while M22 heavy chain mimics the interactions of the TSHα chain. The M22–TSHR complex contains a greater number of hydrogen bonds and salt bridges and fewer hydrophobic interactions than the TSH–TSHR complex, consistent with a higher M22 binding affinity. Furthermore, the surface area formed by TSHR residues N208, Q235, R255, and N256 has been identified as a candidate target region for small molecules which might selectively block binding of autoantibodies to the TSHR.

scholarly journals Modified Potential Functions Result in Enhanced Predictions of a Protein Complex by All-Atom MD Simulations, Confirming a Step-wise Association Process for Native PPIs

2018 ◽  
Author(s):  
Zhen-lu Li ◽  
Matthias Buck
Keyword(s):  
Electrostatic Interaction ◽  
Protein Interactions ◽  
Protein Complex ◽  
Complex Structure ◽  
Md Simulations ◽  
Potential Functions ◽  
Amino Acid Side Chain ◽  
Native Protein ◽  
Protein Protein Interactions ◽  
Steering Mechanism

ABSTRACTNative protein-protein interactions (PPIs) are the cornerstone for understanding the structure, dynamics and mechanisms of function of protein complexes. In this study, we investigate the association of the SAM domains of the EphA2 receptor and SHIP2 enzyme by performing a combined total of 48 μs all-atom molecular dynamics (MD) simulations. While the native SAM heterodimer is only predicted at a low rate of 6.7% with the original CHARMM36 force field, the yield is increased to 16.7% and to 18.3% by scaling the vdW solute-solvent interactions (better fitting the solvation free energy of amino acid side chain analogues) and by an increase of vdW radius of guanidinium interactions, and thus a dramatic reduction of electrostatic interaction between Arg and Glu/Asn in CHARMM36m, respectively. These modifications effectively improve the overly sticky association of proteins, such as ubiquitin, using the original potential function. By analyzing the 25 native SAM complexes formed in the simulations, we find that their formation involves a pre-orientation guided by electrostatic interaction, consistent with an electrostatic steering mechanism. The complex could then transform to the native protein interaction surfaces directly from a well pre-orientated position (Δinterface-RMSD < 5Å). In other cases, modest (< 90°) orientational and/or translational adjustments are needed (5 Å <Δi-RMSD <10 Å) to the native complex. Although the tendency for non-native complexes to dissociate has nearly doubled with the modified potential functions, a re-association to the correct complex structure is still rare. Instead a most non-native complexes are undergoing configurational changes/surface searching, which do not lead to native structures on a timescale of 250 ns. These observations provide a rich picture on mechanisms of protein-protein complex formation, and suggest that computational predictions of native complex protein-protein interactions could be improved further.

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