scholarly journals MLL1 minimal catalytic complex is a dynamic conformational ensemble susceptible to pharmacological allosteric disruption

2018 ◽  
Author(s):  
Lilia Kaustov ◽  
Alexander Lemak ◽  
Hong Wu ◽  
Marco Faini ◽  
Scott Houliston ◽  
...  
Keyword(s):  
Hybrid Methods ◽  
Structural Basis ◽  
Epigenetic Mark ◽  
Conformational Ensemble ◽  
Interaction Sites ◽  
Catalytic Function ◽  
Individual Interaction ◽  
Mixed Lineage Leukaemia ◽  
Repeat Domain ◽  
Histone H3k4

ABSTRACTHistone H3K4 methylation is an epigenetic mark associated with actively transcribed genes. This modification is catalyzed by the mixed lineage leukaemia (MLL) family of histone methyltransferases including MLL1, MLL2, MLL3, MLL4, SET1A and SET1B. Catalytic activity of MLL proteins is dependent on interactions with additional conserved proteins but the structural basis for subunit assembly and the mechanism of regulation is not well understood. We used a hybrid methods approach to study the assembly and biochemical function of the minimally active MLL1 complex (MLL1, WDR5 and RbBP5). A combination of small angle X-ray scattering (SAXS), cross-linking mass spectrometry (XL-MS), NMR spectroscopy, and computational modeling were used to generate a dynamic ensemble model in which subunits are assembled via multiple weak interaction sites. We identified a new interaction site between the MLL1 SET domain and the WD40 repeat domain of RbBP5, and demonstrate the susceptibility of the catalytic function of the complex to disruption of individual interaction sites.

scholarly journals The MLL1 trimeric catalytic complex is a dynamic conformational ensemble stabilized by multiple weak interactions

2019 ◽  
Author(s):  
Lilia Kaustov ◽  
Alexander Lemak ◽  
Hong Wu ◽  
Marco Faini ◽  
Lixin Fan ◽  
...  
Keyword(s):  
Hybrid Methods ◽  
Weak Interactions ◽  
Structural Basis ◽  
Resonance Spectroscopy ◽  
Epigenetic Mark ◽  
Conformational Ensemble ◽  
Interaction Sites ◽  
Catalytic Function ◽  
Mixed Lineage Leukaemia ◽  
Histone H3k4

Abstract Histone H3K4 methylation is an epigenetic mark associated with actively transcribed genes. This modification is catalyzed by the mixed lineage leukaemia (MLL) family of histone methyltransferases including MLL1, MLL2, MLL3, MLL4, SET1A and SET1B. The catalytic activity of this family is dependent on interactions with additional conserved proteins, but the structural basis for subunit assembly and the mechanism of regulation is not well understood. We used a hybrid methods approach to study the assembly and biochemical function of the minimally active MLL1 complex (MLL1, WDR5 and RbBP5). A combination of small angle X-ray scattering, cross-linking mass spectrometry, nuclear magnetic resonance spectroscopy and computational modeling were used to generate a dynamic ensemble model in which subunits are assembled via multiple weak interaction sites. We identified a new interaction site between the MLL1 SET domain and the WD40 β-propeller domain of RbBP5, and demonstrate the susceptibility of the catalytic function of the complex to disruption of individual interaction sites.

scholarly journals Assembly and cryo-EM structures of RNA-specific measles virus nucleocapsids provide mechanistic insight into paramyxoviral replication

2019 ◽  
Vol 116 (10) ◽  
pp. 4256-4264 ◽  
Author(s):  
Ambroise Desfosses ◽  
Sigrid Milles ◽  
Malene Ringkjøbing Jensen ◽  
Serafima Guseva ◽  
Jacques-Philippe Colletier ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Measles Virus ◽  
Rna Binding ◽  
Structural Basis ◽  
Particle Assembly ◽  
Interaction Sites ◽  
Assembly Kinetics ◽  
High Resolution Structure ◽  
Binding Groove

Assembly of paramyxoviral nucleocapsids on the RNA genome is an essential step in the viral cycle. The structural basis of this process has remained obscure due to the inability to control encapsidation. We used a recently developed approach to assemble measles virus nucleocapsid-like particles on specific sequences of RNA hexamers (poly-Adenine and viral genomic 5′) in vitro, and determined their cryoelectron microscopy maps to 3.3-Å resolution. The structures unambiguously determine 5′ and 3′ binding sites and thereby the binding-register of viral genomic RNA within nucleocapsids. This observation reveals that the 3′ end of the genome is largely exposed in fully assembled measles nucleocapsids. In particular, the final three nucleotides of the genome are rendered accessible to the RNA-dependent RNA polymerase complex, possibly enabling efficient RNA processing. The structures also reveal local and global conformational changes in the nucleoprotein upon assembly, in particular involving helix α6 and helix α13 that form edges of the RNA binding groove. Disorder is observed in the bound RNA, localized at one of the two backbone conformational switch sites. The high-resolution structure allowed us to identify putative nucleobase interaction sites in the RNA-binding groove, whose impact on assembly kinetics was measured using real-time NMR. Mutation of one of these sites, R195, whose sidechain stabilizes both backbone and base of a bound nucleic acid, is thereby shown to be essential for nucleocapsid-like particle assembly.

scholarly journals Targeting Intrinsically Disordered Proteins through Dynamic Interactions

Biomolecules ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 743
Author(s):  
Jianlin Chen ◽  
Xiaorong Liu ◽  
Jianhan Chen
Keyword(s):  
Small Molecules ◽  
Intrinsically Disordered Proteins ◽  
Gpu Computing ◽  
Disordered Proteins ◽  
Atomistic Simulations ◽  
Conformational Ensemble ◽  
Processing Unit ◽  
Dynamic Interactions ◽  
Intrinsically Disordered ◽  
Interaction Sites

Intrinsically disordered proteins (IDPs) are over-represented in major disease pathways and have attracted significant interest in understanding if and how they may be targeted using small molecules for therapeutic purposes. While most existing studies have focused on extending the traditional structure-centric drug design strategies and emphasized exploring pre-existing structure features of IDPs for specific binding, several examples have also emerged to suggest that small molecules could achieve specificity in binding IDPs and affect their function through dynamic and transient interactions. These dynamic interactions can modulate the disordered conformational ensemble and often lead to modest compaction to shield functionally important interaction sites. Much work remains to be done on further elucidation of the molecular basis of the dynamic small molecule–IDP interaction and determining how it can be exploited for targeting IDPs in practice. These efforts will rely critically on an integrated experimental and computational framework for disordered protein ensemble characterization. In particular, exciting advances have been made in recent years in enhanced sampling techniques, Graphic Processing Unit (GPU)-computing, and protein force field optimization, which have now allowed rigorous physics-based atomistic simulations to generate reliable structure ensembles for nontrivial IDPs of modest sizes. Such de novo atomistic simulations will play crucial roles in exploring the exciting opportunity of targeting IDPs through dynamic interactions.

Structural Basis for Ca2+-regulated Muscle Relaxation at Interaction Sites of Troponin with Actin and Tropomyosin

2005 ◽  
Vol 352 (1) ◽  
pp. 178-201 ◽  
Author(s):  
Kenji Murakami ◽  
Fumiaki Yumoto ◽  
Shin-ya Ohki ◽  
Takuo Yasunaga ◽  
Masaru Tanokura ◽  
...  
Keyword(s):  
Muscle Relaxation ◽  
Structural Basis ◽  
Interaction Sites

scholarly journals Mapping the conformational landscape of a dynamic enzyme by multitemperature and XFEL crystallography

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Daniel A Keedy ◽  
Lillian R Kenner ◽  
Matthew Warkentin ◽  
Rahel A Woldeyes ◽  
Jesse B Hopkins ◽  
...  
Keyword(s):  
Active Site ◽  
Protein Function ◽  
Conformational Ensemble ◽  
Conformational Heterogeneity ◽  
Time Resolved ◽  
Protein Motions ◽  
Catalytic Function ◽  
Temperature Dependences ◽  
Site Network ◽  
Response To Temperature

Determining the interconverting conformations of dynamic proteins in atomic detail is a major challenge for structural biology. Conformational heterogeneity in the active site of the dynamic enzyme cyclophilin A (CypA) has been previously linked to its catalytic function, but the extent to which the different conformations of these residues are correlated is unclear. Here we compare the conformational ensembles of CypA by multitemperature synchrotron crystallography and fixed-target X-ray free-electron laser (XFEL) crystallography. The diffraction-before-destruction nature of XFEL experiments provides a radiation-damage-free view of the functionally important alternative conformations of CypA, confirming earlier synchrotron-based results. We monitored the temperature dependences of these alternative conformations with eight synchrotron datasets spanning 100-310 K. Multiconformer models show that many alternative conformations in CypA are populated only at 240 K and above, yet others remain populated or become populated at 180 K and below. These results point to a complex evolution of conformational heterogeneity between 180-–240 K that involves both thermal deactivation and solvent-driven arrest of protein motions in the crystal. The lack of a single shared conformational response to temperature within the dynamic active-site network provides evidence for a conformation shuffling model, in which exchange between rotamer states of a large aromatic ring in the middle of the network shifts the conformational ensemble for the other residues in the network. Together, our multitemperature analyses and XFEL data motivate a new generation of temperature- and time-resolved experiments to structurally characterize the dynamic underpinnings of protein function.

scholarly journals The molecular mechanism of Hsp90-induced oligomerization of Tau

2019 ◽  
Author(s):  
Sabrina Weickert ◽  
Magdalena Wawrzyniuk ◽  
Laura John ◽  
Stefan G. D. Rüdiger ◽  
Malte Drescher
Keyword(s):  
Structural Information ◽  
Conformational Ensemble ◽  
Toxic Agent ◽  
Paramagnetic Resonance ◽  
Biologically Relevant ◽  
Intrinsically Disordered ◽  
Tau Oligomers ◽  
Paper Clip ◽  
Repeat Domain ◽  
Electron Paramagnetic

AbstractAggregation of the microtubule-associated protein Tau is a hallmark of Alzheimer’s disease with Tau oligomers suspected as the most toxic agent. Tau is a client of Hsp90, though it is unclear whether and how the chaperone massages the structure of intrinsically disordered Tau. Using electron paramagnetic resonance, we extract structural information from the very broad conformational ensemble of Tau: Tau in solution is highly dynamic and polymorphic, though ‘paper-clip’-shaped by long-range contacts. Interaction with Hsp90 promotes an open Tau conformation, which we identify as the molecular basis for the formation of small Tau oligomers by exposure of the aggregation-prone repeat domain to other Tau molecules. At the same time, formation of Tau fibrils is inhibited. We therefore provide the nanometer-scale zoom into chaperoning an amyloid client, highlighting formation of oligomers as the consequence of this biologically relevant interaction.

scholarly journals Structural basis for snRNA recognition by the double-WD40 repeat domain of Gemin5

2016 ◽  
Vol 30 (21) ◽  
pp. 2391-2403 ◽  
Author(s):  
Wenxing Jin ◽  
Yi Wang ◽  
Chao-Pei Liu ◽  
Na Yang ◽  
Mingliang Jin ◽  
...  
Keyword(s):  
Structural Basis ◽  
Wd40 Repeat ◽  
Repeat Domain

Structural basis for histone H3K4me3 recognition by the N-terminal domain of the PHD finger protein Spp1

2019 ◽  
Vol 476 (13) ◽  
pp. 1957-1973 ◽  
Author(s):  
Chao He ◽  
Ning Liu ◽  
Dongya Xie ◽  
Yanhong Liu ◽  
Yazhong Xiao ◽  
...  
Keyword(s):  
Strand Break ◽  
Structural Basis ◽  
Titration Calorimetry ◽  
Chromatin Binding ◽  
Phd Finger ◽  
Terminal Domain ◽  
Saccharomyces Pombe ◽  
Plant Homeodomain ◽  
Histone H3k4

Abstract Saccharomyces cerevisiae Spp1, a plant homeodomain (PHD) finger containing protein, is a critical subunit of the histone H3K4 methyltransferase complex of proteins associated with Set1 (COMPASS). The chromatin binding affinity of the PHD finger of Spp1 has been proposed to modulate COMPASS activity. During meiosis, Spp1 plays another role in promoting programmed double-strand break (DSB) formation by binding H3K4me3 via its PHD finger and interacting with a DSB protein, Mer2. However, how the Spp1 PHD finger performs site-specific readout of H3K4me3 is still not fully understood. In the present study, we determined the crystal structure of the highly conserved Spp1 N-terminal domain (Sc_Spp1NTD) in complex with the H3K4me3 peptide. The structure shows that Sc_Spp1NTD comprises a PHD finger responsible for methylated H3K4 recognition and a C3H-type zinc finger necessary to ensure the overall structural stability. Our isothermal titration calorimetry results show that binding of H3K4me3 to Sc_Spp1NTD is mildly inhibited by H3R2 methylation, weakened by H3T6 phosphorylation, and abrogated by H3T3 phosphorylation. This histone modification cross-talk, which is conserved in the Saccharomyces pombe and mammalian orthologs of Sc_Spp1 in vitro, can be rationalized structurally and might contribute to the roles of Spp1 in COMPASS activity regulation and meiotic recombination.

scholarly journals Structural Basis of Ligand Interactions of the Large Extracellular Domain of Tetraspanin CD81

2012 ◽  
Vol 86 (18) ◽  
pp. 9606-9616 ◽  
Author(s):  
Sundaresan Rajesh ◽  
Pooja Sridhar ◽  
Birke Andrea Tews ◽  
Lucie Fénéant ◽  
Laurence Cocquerel ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Primary Liver Cancer ◽  
Structural Basis ◽  
Extracellular Loop ◽  
Entry Level ◽  
Interaction Sites ◽  
Loop Region ◽  
Binding Interface ◽  
Ligand Interactions ◽  
Large Extracellular Loop

Hepatitis C virus (HCV) causes chronic liver disease, cirrhosis, and primary liver cancer. Despite 130 million people being at risk worldwide, no vaccine exists, and effective therapy is limited by drug resistance, toxicity, and high costs. The tetraspanin CD81 is an essential entry-level receptor required for HCV infection of hepatocytes and represents a critical target for intervention. In this study, we report the first structural characterization of the large extracellular loop of CD81, expressed in mammalian cells and studied in physiological solutions. The HCV E2 glycoprotein recognizes CD81 through a dynamic loop on the helical bundle, which was shown by nuclear magnetic resonance (NMR) spectroscopy to adopt a conformation distinct from that seen in crystals. A novel membrane binding interface was revealed adjacent to the exposed HCV interaction site in the extracellular loop of CD81. The binding pockets for two proposed inhibitors of the CD81-HCV interaction, namely, benzyl salicylate and fexofenadine, were shown to overlap the HCV and membrane interaction sites. Although the dynamic loop region targeted by these compounds presents challenges for structure-based design, the NMR assignments enable realistic screening and validation of ligands. Together, these data provide an improved avenue for developing potent agents that specifically block CD81-HCV interaction and also pave a way for elucidating the recognition mechanisms of diverse tetraspanins.

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