scholarly journals Structural evolutionary analysis predicts functional sites in the artemisinin resistance malaria protein K13

2018 ◽  
Author(s):  
Romain Coppée ◽  
Daniel C. Jeffares ◽  
Audrey Sabbagh ◽  
Jérôme Clain
Keyword(s):  
Structural Information ◽  
Artemisinin Resistance ◽  
Evolutionary Analysis ◽  
Functional Sites ◽  
Btb Domain ◽  
Binding Surface ◽  
Species Evolution ◽  
Binding Residues ◽  
Falciparum Protein

AbstractK13 is an essential Plasmodium falciparum protein that plays a key role in malaria resistance to artemisinins. Although K13 resembles BTB- and Kelch/propeller-containing proteins involved in ubiquitin ligase complexes, its functional sites remain uncharacterized. Using evolutionary and structural information, we searched for the most conserved K13 sites across Apicomplexa species evolution to identify sub-regions of K13 that are likely functional. An amino acid electropositive ‘patch’ in the K13 propeller domain has a dense concentration of extraordinarily conserved positions located at a shallow pocket, suggesting a role as binding surface. When applied to experimentally-characterized BTB-Kelch proteins, our strategy successfully identifies the validated substrate-binding residues within their own propeller shallow pocket. Another patch of slowly evolving sites is identified in the K13 BTB domain which partially overlaps the surface that binds to Cullin proteins in BTB-Cullin complexes. We provide candidate binding sites in K13 propeller and BTB domains for functional follow-up studies.

scholarly journals Functional Importance of Hydrophobic Patches on the Ebola Virus VP35 IFN-Inhibitory Domain

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2316
Author(s):  
Nodoka Kasajima ◽  
Keita Matsuno ◽  
Hiroko Miyamoto ◽  
Masahiro Kajihara ◽  
Manabu Igarashi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ebola Virus ◽  
Structural Information ◽  
Site Directed Mutagenesis ◽  
Genome Replication ◽  
Type I ◽  
Multifunctional Protein ◽  
Functional Sites ◽  
Inhibitory Function ◽  
Inhibitory Domain

Viral protein 35 (VP35) of Ebola virus (EBOV) is a multifunctional protein that mainly acts as a viral polymerase cofactor and an interferon antagonist. VP35 interacts with the viral nucleoprotein (NP) and double-stranded RNA for viral RNA transcription/replication and inhibition of type I interferon (IFN) production, respectively. The C-terminal portion of VP35, which is termed the IFN-inhibitory domain (IID), is important for both functions. To further identify critical regions in this domain, we analyzed the physical properties of the surface of VP35 IID, focusing on hydrophobic patches, which are expected to be functional sites that are involved in interactions with other molecules. Based on the known structural information of VP35 IID, three hydrophobic patches were identified on its surface and their biological importance was investigated using minigenome and IFN-β promoter-reporter assays. Site-directed mutagenesis revealed that some of the amino acid substitutions that were predicted to disrupt the hydrophobicity of the patches significantly decreased the efficiency of viral genome replication/transcription due to reduced interaction with NP, suggesting that the hydrophobic patches might be critical for the formation of a replication complex through the interaction with NP. It was also found that the hydrophobic patches were involved in the IFN-inhibitory function of VP35. These results highlight the importance of hydrophobic patches on the surface of EBOV VP35 IID and also indicate that patch analysis is useful for the identification of amino acid residues that directly contribute to protein functions.

scholarly journals Wxlv, the Ancestral Allele of Rice Waxy Gene

Proceedings ◽  
2020 ◽  
Vol 36 (1) ◽  
pp. 140 ◽  
Author(s):  
Changquan Zhang ◽  
Jihui Zhu ◽  
Shengjie Chen ◽  
Qiaoquan Liu
Keyword(s):  
Amylose Content ◽  
Waxy Gene ◽  
Evolutionary Analysis ◽  
Ancestral Allele ◽  
Cultivated Rice ◽  
Functional Sites ◽  
Wx Locus ◽  
Wx Gene ◽  
Shed Light

In rice endosperms, the Waxy (Wx) gene is important for amylose synthesis, and various Wx alleles control the amylose content and affect the taste of cooked rice. Herein, we report the cloning of the ancestral allele Wxlv of the Wx locus, which affects the mouthfeel of rice grains by modulating the size of amylose molecules. Using evolutionary analysis, we demonstrated that Wxlv originated directly from wild rice, and the three major Wx alleles in cultivated rice (Wxb, Wxa, and Wxin) differentiated after the substitution of one base pair at the functional sites. These data indicate that the Wxlv allele played an important role in artificial selection and domestication. The findings also shed light on the evolution of various Wx alleles, which have greatly contributed to improving the eating and cooking quality of rice.

NMR for the design of functional mimetics of protein-protein interactions: one key is in the building of bridges

1998 ◽  
Vol 76 (2-3) ◽  
pp. 177-188 ◽  
Author(s):  
Jianxing Song ◽  
Feng Ni
Keyword(s):  
Protein Interactions ◽  
Binding Sites ◽  
Experimental Approach ◽  
Structural Information ◽  
Peptide Ligands ◽  
Protein Protein Interactions ◽  
Binding Residues ◽  
Related Proteins ◽  
Binding Inhibitors

Using the design of bivalent and bridge-binding inhibitors of thrombin as an example, we review an NMR-based experimental approach for the design of functional mimetics of protein-protein interactions. The strategy includes: (i) identification of binding residues in peptide ligands by differential resonance perturbation, (ii) determination of protein-bound structures of peptide ligands by use of transferred NOEs, (iii) minimization of larger protein and peptide ligands on the basis of NMR structural information, and (iv) linkage of two weakly binding mimetics to produce an inhibitor with enhanced affinity and specificity. This approach can be especially effective for the design of potent and selective functional mimetics of protein-protein interactions because it is less likely that the surfaces of two related proteins or enzymes share two identical binding sites or regions.Key words: NMR, protein-protein interactions, functional mimetics, bridge-binding inhibitors, thrombin.

scholarly journals Integrated structural and evolutionary analysis reveals common mechanisms underlying adaptive evolution in mammals

2020 ◽  
Vol 117 (11) ◽  
pp. 5977-5986 ◽  
Author(s):  
Greg Slodkowicz ◽  
Nick Goldman
Keyword(s):  
Positive Selection ◽  
Adaptive Evolution ◽  
Evolutionary Biology ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Protein Structures ◽  
Unexpected Finding ◽  
Evolutionary Analysis ◽  
Genomic Scale ◽  
Evolution Of Mammals

Understanding the molecular basis of adaptation to the environment is a central question in evolutionary biology, yet linking detected signatures of positive selection to molecular mechanisms remains challenging. Here we demonstrate that combining sequence-based phylogenetic methods with structural information assists in making such mechanistic interpretations on a genomic scale. Our integrative analysis shows that positively selected sites tend to colocalize on protein structures and that positively selected clusters are found in functionally important regions of proteins, indicating that positive selection can contravene the well-known principle of evolutionary conservation of functionally important regions. This unexpected finding, along with our discovery that positive selection acts on structural clusters, opens previously unexplored strategies for the development of better models of protein evolution. Remarkably, proteins where we detect the strongest evidence of clustering belong to just two functional groups: Components of immune response and metabolic enzymes. This gives a coherent picture of pathogens and xenobiotics as important drivers of adaptive evolution of mammals.

Micro- and Cryo-Techniques Prevent the Loss of Structural Information. EM-Studies on High-Pressure Frozen Tissues, Suspensions and Cell Monolayer.

1999 ◽  
Vol 5 (S2) ◽  
pp. 430-431
Author(s):  
H. Hohenberg
Keyword(s):  
Electron Microscopy ◽  
High Pressure ◽  
Structural Information ◽  
Cell Monolayer ◽  
Freeze Substitution ◽  
Fine Needle Biopsy ◽  
Specimen Preparation ◽  
Optical Instruments ◽  
Cultivation Technique

Cells are information driven systems. Cellular information is stored in certain molecules, at certain places, in a certain concentration, at a particular time and under given physiological conditions. The goal of biological electron microscopy is to provide this information network, to correlate the cellular ultrastructure and its function. In this sense, it is essential to combine the high resolution of our electron optical instruments with a high information density of the biological system. Most of the structural information is lost in the course of the different preparation steps prior to electron microscopy. For this reason it is necessary that all preparation steps such as: 1. sampling: e.g. excision of tissues, 2. cryoimmobilisation, 3. follow-up procedures: e.g. freeze-fracturing, freeze-substitution and embedding, should have identical high quality levels preventing or minimizing the loss of structural information. To this aim our methodological activities focus on the development of special micro-techniques for the sampling of: a) native tissues, with an automatic fine-needle biopsy technique (1), of b) suspensions, with a special cellulose capillary technique (2), of c) cell monolayer, with a thin film cultivation technique (3) and the application/perfection of cryotechniques (high-pressure freezing (HPF) and freeze-substitution). In particular, the high-pressure freezer (HPM 010, Bal-Tec) has proven to be a highly useful tool for successful cryoimmobilization of almost any kinds of cells and tissues, bulk specimens (< 200 μm in thickness) being included. This freezing technique does not require any cryoprotection, and if combined with our micro-techniques the risk of inducing artefacts as a result of specimen preparation prior to freezing is minimized.

Significance of panendoscopy and CT in the follow-up and management of squamous cell carcinoma of the head and neck: A retrospective clinical assessment.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e17003-e17003
Author(s):  
Adrian Muenscher ◽  
Susanne Sehner ◽  
Jegane Taleh ◽  
Silke Tribius ◽  
Carsten Dalchow ◽  
...  
Keyword(s):  
Head And Neck ◽  
Squamous Cell ◽  
Structural Information ◽  
Clinical Investigation ◽  
Medical Center ◽  
Tnm Staging ◽  
Histological Evaluation ◽  
Female Ratio ◽  
Male Predominance

e17003 Background: Treatment of patients with head and neck squamous cell carcinomas (HNSCC) require an evidence-based multidisciplinary approach which varies from medical centres. Panendoscopy has been traditionally used as a routine screening examination tool. We discuss and compare its clinical effectiveness, safety and predictive value in patients with computed tomography (CT) scans in a routine outpatient follow up. Methods: Between 2004-2007, in a retrospective study (204 patients; male:156 (76.5%); female:48 (23.5%))with head and neck cancer, fulfilled all inclusion and exclusion criteria were evaluated at the University Medical Center Hamburg-Eppendorf, Germany. Follow-ups were performed every 2 to 3 months for the first 3 years and every 6 month for the following 2 years. UICC-TNM staging system divided patients into seven categories. Results: The mean age of patients was (male: 59.2±9.3 years; female: 59.5±10.2 years), the highest incidence rate was recorded in the 51 to 60 age group (n = 77). There was a male predominance, with a male/female ratio of 3:1. In comparing normal versus abnormal CT a 12-fold higher frequency of recurrence (odds ratio [OR]; 95% CI: 3.99, 37.51; p<0.001) was interpreted.Additionally, patient anamnesis (OR: 3.33; 95% CI: 1.57, 7.07; p=0.002), clinical investigation (OR: 5.85; 95% CI: 3.01,11.36; p<0.001) and recurrence pattern (OR: 4.82; 95% CI: 2.26,10.32; p<0.001) was observed. Conclusions: In staging / follow-up HNSCC patients, CT-diagnostics also provided similar levels of accuracy as panendoscopy. Panendoscopy in patients with HNSCC was avoided (169 out of 364) when the diagnostic parameters remained satisfactory. Higher patient satisfaction, structural information at a high spatial resolution and comfort was noted in the CT group, indicating a gradual move towards CT diagnostics. For safety-related reasons in particular, we recommend panendoscopy for histological evaluation i.e.in predicting cancer recurrence and progression.

scholarly journals Proteolysis of Monomeric Recombinant Rotavirus VP4 Yields an Oligomeric VP5* Core

2001 ◽  
Vol 75 (16) ◽  
pp. 7339-7350 ◽  
Author(s):  
Philip R. Dormitzer ◽  
Harry B. Greenberg ◽  
Stephen C. Harrison
Keyword(s):  
Analytical Ultracentrifugation ◽  
Structural Information ◽  
Sodium Dodecyl ◽  
Heptad Repeat ◽  
Membrane Penetration ◽  
Electron Cryomicroscopy ◽  
Infected Insect ◽  
Binding Residues ◽  
Sequential Digestion ◽  
Outer Capsid

ABSTRACT Rotavirus particles are activated for cell entry by trypsin cleavage of the outer capsid spike protein, VP4, into a hemagglutinin, VP8*, and a membrane penetration protein, VP5*. We have purified rhesus rotavirus VP4, expressed in baculovirus-infected insect cells. Purified VP4 is a soluble, elongated monomer, as determined by analytical ultracentrifugation. Trypsin cleaves purified VP4 at a number of sites that are protected on the virion and yields a heterogeneous group of protease-resistant cores of VP5*. The most abundant tryptic VP5* core is trimmed past the N terminus associated with activation for virus entry into cells. Sequential digestion of purified VP4 with chymotrypsin and trypsin generates homogeneous VP8* and VP5* cores (VP8CT and VP5CT, respectively), which have the authentic trypsin cleavages in the activation region. VP8CT is a soluble monomer composed primarily of β-sheets. VP5CT forms sodium dodecyl sulfate-resistant dimers. These results suggest that trypsinization of rotavirus particles triggers a rearrangement in the VP5* region of VP4 to yield the dimeric spikes observed in icosahedral image reconstructions from electron cryomicroscopy of trypsinized rotavirus virions. The solubility of VP5CT and of trypsinized rotavirus particles suggests that the trypsin-triggered conformational change primes VP4 for a subsequent rearrangement that accomplishes membrane penetration. The domains of VP4 defined by protease analysis contain all mapped neutralizing epitopes, sialic acid binding residues, the heptad repeat region, and the membrane permeabilization region. This biochemical analysis of VP4 provides sequence-specific structural information that complements electron cryomicroscopy data and defines targets and strategies for atomic-resolution structural studies.

scholarly journals Rapid molecular evolution across amniotes of the IIS/TOR network

2015 ◽  
Vol 112 (22) ◽  
pp. 7055-7060 ◽  
Author(s):  
Suzanne E. McGaugh ◽  
Anne M. Bronikowski ◽  
Chih-Horng Kuo ◽  
Dawn M. Reding ◽  
Elizabeth A. Addis ◽  
...  
Keyword(s):  
Molecular Evolution ◽  
Positive Selection ◽  
Insulin Receptor ◽  
Metabolic Diseases ◽  
Vital Role ◽  
Evolutionary Rates ◽  
Evolutionary Analysis ◽  
Binding Surface ◽  
Tor Network ◽  
Positively Selected Sites

The insulin/insulin-like signaling and target of rapamycin (IIS/TOR) network regulates lifespan and reproduction, as well as metabolic diseases, cancer, and aging. Despite its vital role in health, comparative analyses of IIS/TOR have been limited to invertebrates and mammals. We conducted an extensive evolutionary analysis of the IIS/TOR network across 66 amniotes with 18 newly generated transcriptomes from nonavian reptiles and additional available genomes/transcriptomes. We uncovered rapid and extensive molecular evolution between reptiles (including birds) and mammals: (i) the IIS/TOR network, including the critical nodes insulin receptor substrate (IRS) and phosphatidylinositol 3-kinase (PI3K), exhibit divergent evolutionary rates between reptiles and mammals; (ii) compared with a proxy for the rest of the genome, genes of the IIS/TOR extracellular network exhibit exceptionally fast evolutionary rates; and (iii) signatures of positive selection and coevolution of the extracellular network suggest reptile- and mammal-specific interactions between members of the network. In reptiles, positively selected sites cluster on the binding surfaces of insulin-like growth factor 1 (IGF1), IGF1 receptor (IGF1R), and insulin receptor (INSR); whereas in mammals, positively selected sites clustered on the IGF2 binding surface, suggesting that these hormone-receptor binding affinities are targets of positive selection. Further, contrary to reports that IGF2R binds IGF2 only in marsupial and placental mammals, we found positively selected sites clustered on the hormone binding surface of reptile IGF2R that suggest that IGF2R binds to IGF hormones in diverse taxa and may have evolved in reptiles. These data suggest that key IIS/TOR paralogs have sub- or neofunctionalized between mammals and reptiles and that this network may underlie fundamental life history and physiological differences between these amniote sister clades.

scholarly journals AlphaFold-aware prediction of protein-DNA binding sites using graph transformer

2021 ◽  
Author(s):  
Qianmu Yuan ◽  
Sheng Chen ◽  
Jiahua Rao ◽  
Shuangjia Zheng ◽  
Huiying Zhao ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Sites ◽  
Structure Prediction ◽  
Spatial Information ◽  
Structural Information ◽  
Biological Activities ◽  
Protein Structures ◽  
Dna Binding Sites ◽  
Novel Drugs ◽  
Binding Residues

AbstractMotivationProtein-DNA interactions play crucial roles in the biological systems, and identifying protein-DNA binding sites is the first step for mechanistic understanding of various biological activities (such as transcription and repair) and designing novel drugs. How to accurately identify DNA-binding residues from only protein sequence remains a challenging task. Currently, most existing sequence-based methods only consider contextual features of the sequential neighbors, which are limited to capture spatial information.ResultsBased on the recent breakthrough in protein structure prediction by AlphaFold2, we propose an accurate predictor, GraphSite, for identifying DNA-binding residues based on the structural models predicted by AlphaFold2. Here, we convert the binding site prediction problem into a graph node classification task and employ a transformerbased variant model to take the protein structural information into account. By leveraging predicted protein structures and graph transformer, GraphSite substantially improves over the latest sequence-based and structure-based methods. The algorithm was further confirmed on the independent test set of 196 proteins, where GraphSite surpasses the state-of-the-art structure-based method by 12.3% in AUPR and 9.3% in MCC, [email protected]

scholarly journals Comparative structural and evolutionary analyses predict functional sites in the artemisinin resistance malaria protein K13

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Romain Coppée ◽  
Daniel C. Jeffares ◽  
Maria A. Miteva ◽  
Audrey Sabbagh ◽  
Jérôme Clain
Keyword(s):  
Artemisinin Resistance ◽  
Functional Sites
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