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 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.

scholarly journals Short loop functional commonality identified in leukaemia proteome highlights crucial protein sub-networks

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Sun Sook Chung ◽  
Joseph C F Ng ◽  
Anna Laddach ◽  
N Shaun B Thomas ◽  
Franca Fraternali
Keyword(s):  
Protein Interactions ◽  
Large Scale ◽  
Interaction Network ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Ppi Networks ◽  
Short Loop ◽  
New Strategy ◽  
Loop Network ◽  
Protein Protein Interaction Network

Abstract Direct drug targeting of mutated proteins in cancer is not always possible and efficacy can be nullified by compensating protein–protein interactions (PPIs). Here, we establish an in silico pipeline to identify specific PPI sub-networks containing mutated proteins as potential targets, which we apply to mutation data of four different leukaemias. Our method is based on extracting cyclic interactions of a small number of proteins topologically and functionally linked in the Protein–Protein Interaction Network (PPIN), which we call short loop network motifs (SLM). We uncover a new property of PPINs named ‘short loop commonality’ to measure indirect PPIs occurring via common SLM interactions. This detects ‘modules’ of PPI networks enriched with annotated biological functions of proteins containing mutation hotspots, exemplified by FLT3 and other receptor tyrosine kinase proteins. We further identify functional dependency or mutual exclusivity of short loop commonality pairs in large-scale cellular CRISPR–Cas9 knockout screening data. Our pipeline provides a new strategy for identifying new therapeutic targets for drug discovery.

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 Mitochondria: Regulators of Cell Death and Survival

2002 ◽  
Vol 2 ◽  
pp. 1569-1578 ◽  
Author(s):  
David J. Granville ◽  
Roberta A. Gottlieb
Keyword(s):  
Cell Death ◽  
Conformational Changes ◽  
Protein Interactions ◽  
Apoptotic Cell ◽  
Critical Role ◽  
Death Process ◽  
Protein Protein Interactions ◽  
Ionic Changes ◽  
A Cell ◽  
Cyt C

The past 5 years has seen an intense surge in research devoted toward understanding the critical role of mitochondria in the regulation of cell death. Apoptosis can be initiated by a wide array of stimuli, inducing multiple signaling pathways that, for the most part, converge at the mitochondrion. Although classically considered the powerhouses of the cell, it is now understood that mitochondria are also “gatekeepers” that ultimately determine the fate of the cell. The mitochondrial decision as to whether a cell lives or dies is complex, involving protein-protein interactions, ionic changes, reactive oxygen species, and other mechanisms that require further elucidation. Once the death process is initiated, mitochondria undergo conformational changes, resulting in the release of cytochrome c (cyt c), caspases, endonucleases, and other factors leading to the onset and execution of apoptosis. The present review attempts to outline the complex milieu of events regulating the mitochondrial commitment to and processes involved in the implementation of the executioner phase of apoptotic cell death.

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.

scholarly journals Molecular basis for the interaction between human choline kinase alpha and the SH3 domain of the c-Src tyrosine kinase

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Stefanie L. Kall ◽  
Kindra Whitlatch ◽  
Thomas E. Smithgall ◽  
Arnon Lavie
Keyword(s):  
Crystal Structures ◽  
Protein Interactions ◽  
Catalytic Domain ◽  
Sh3 Domain ◽  
Crystallographic Analysis ◽  
Choline Kinase ◽  
Protein Protein Interactions ◽  
Src Tyrosine Kinase ◽  
Patient Prognosis ◽  
Co Precipitation

AbstractCholine kinase alpha is a 457-residue protein that catalyzes the reaction between ATP and choline to yield ADP and phosphocholine. This metabolic action has been well studied because of choline kinase’s link to cancer malignancy and poor patient prognosis. As the myriad of x-ray crystal structures available for this enzyme show, chemotherapeutic drug design has centered on stopping the catalytic activity of choline kinase and reducing the downstream metabolites it produces. Furthermore, these crystal structures only reveal the catalytic domain of the protein, residues 80–457. However, recent studies provide evidence for a non-catalytic protein-binding role for choline kinase alpha. Here, we show that choline kinase alpha interacts with the SH3 domain of c-Src. Co-precipitation assays, surface plasmon resonance, and crystallographic analysis of a 1.5 Å structure demonstrate that this interaction is specific and is mediated by the poly-proline region found N-terminal to the catalytic domain of choline kinase. Taken together, these data offer strong evidence that choline kinase alpha has a heretofore underappreciated role in protein-protein interactions, which offers an exciting new way to approach drug development against this cancer-enhancing protein.

Complex structure of the acyltransferase VinK and the carrier protein VinL with a pantetheine cross-linking probe

2021 ◽  
Vol 77 (9) ◽  
pp. 294-302
Author(s):  
Akimasa Miyanaga ◽  
Risako Ouchi ◽  
Fumitaka Kudo ◽  
Tadashi Eguchi
Keyword(s):  
Crystal Structure ◽  
Protein Interactions ◽  
Fatty Acid Biosynthesis ◽  
Structural Information ◽  
Complex Structure ◽  
Cross Linking ◽  
Carrier Protein ◽  
Protein Protein Interactions ◽  
Interaction Interface ◽  
Covalent Complex

Acyltransferases are responsible for the selection and loading of acyl units onto carrier proteins in polyketide and fatty-acid biosynthesis. Despite the importance of protein–protein interactions between the acyltransferase and the carrier protein, structural information on acyltransferase–carrier protein interactions is limited because of the transient interactions between them. In the biosynthesis of the polyketide vicenistatin, the acyltransferase VinK recognizes the carrier protein VinL for the transfer of a dipeptidyl unit. The crystal structure of a VinK–VinL covalent complex formed with a 1,2-bismaleimidoethane cross-linking reagent has been determined previously. Here, the crystal structure of a VinK–VinL covalent complex formed with a pantetheine cross-linking probe is reported at 1.95 Å resolution. In the structure of the VinK–VinL–probe complex, the pantetheine probe that is attached to VinL is covalently connected to the side chain of the mutated Cys106 of VinK. The interaction interface between VinK and VinL is essentially the same in the two VinK–VinL complex structures, although the position of the pantetheine linker slightly differs. This structural observation suggests that interface interactions are not affected by the cross-linking strategy used.

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