Conformational Antibody Binding to a Native, Cell-Free Expressed GPCR in Block Copolymer Membranes
Publication Date
October 20, 2014
Journal
PLOS ONE
Authors
Hans Peter M. De Hoog, Esther M. Lin Jie Rong, Sourabh Banerjee, Fabien M. Décaillot, et al
Volume
9
Issue
10
Pages
e110847
DOI
https://dx.plos.org/10.1371/journal.pone.0110847
Publisher URL
http://journals.plos.org/plosone/article?id=10.1371%2Fjournal.pone.0110847
PubMed
http://www.ncbi.nlm.nih.gov/pubmed/25329156
PubMed Central
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4203850
Europe PMC
http://europepmc.org/abstract/MED/25329156
Web of Science
000343942100100
Scopus
84908200314
Mendeley
http://www.mendeley.com/research/conformational-antibody-binding-native-cellfree-expressed-gpcr-block-copolymer-membranes
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Mendeley | Further Information

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Scopus | Further Information

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/1724376"], "description"=>"<p>A: Ab was injected at increasing concentrations (6.25–400 nM) over 100 s, followed by a buffer wash (without regeneration) between injections (immobilization level: ca. 5000 RU; biotin/streptavidin immobilization). B. Saturation binding of 125-I SDF1α to CXCR4-ACMs. A dissociation constant of 8.4 nM was determined. C. The same series of measurements as shown in Fig. 2 A, conducted using immobilized VLPS (immobilization level: 5000 RU).</p>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209984, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0110847.g002", "stats"=>{"downloads"=>0, "page_views"=>24, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Kinetic_screening_of_12G5_mAb_binding_to_CXCR4_ACMs_immobilized_onto_biosensor_chips_/1209984", "title"=>"Kinetic screening of 12G5 mAb binding to CXCR4-ACMs immobilized onto biosensor chips.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-10-20 02:48:45"}
  • {"files"=>["https://ndownloader.figshare.com/files/1724385", "https://ndownloader.figshare.com/files/1724387", "https://ndownloader.figshare.com/files/1724388", "https://ndownloader.figshare.com/files/1724389"], "description"=>"<div><p>G-protein coupled receptors (GPCRs) play a key role in physiological processes and are attractive drug targets. Their biophysical characterization is, however, highly challenging because of their innate instability outside a stabilizing membrane and the difficulty of finding a suitable expression system. We here show the cell-free expression of a GPCR, CXCR4, and its direct embedding in diblock copolymer membranes. The polymer-stabilized CXCR4 is readily immobilized onto biosensor chips for label-free binding analysis. Kinetic characterization using a conformationally sensitive antibody shows the receptor to exist in the correctly folded conformation, showing binding behaviour that is commensurate with heterologously expressed CXCR4.</p></div>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209993, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0110847.s001", "https://dx.doi.org/10.1371/journal.pone.0110847.s002", "https://dx.doi.org/10.1371/journal.pone.0110847.s003", "https://dx.doi.org/10.1371/journal.pone.0110847.s004"], "stats"=>{"downloads"=>3, "page_views"=>51, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Conformational_Antibody_Binding_to_a_Native_Cell_Free_Expressed_GPCR_in_Block_Copolymer_Membranes_/1209993", "title"=>"Conformational Antibody Binding to a Native, Cell-Free Expressed GPCR in Block Copolymer Membranes", "pos_in_sequence"=>0, "defined_type"=>4, "published_date"=>"2014-10-20 02:48:45"}
  • {"files"=>["https://ndownloader.figshare.com/files/1724382"], "description"=>"[a]<p>KD for 12G5 binding to CXCR4-ACMS shown in <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110847#pone-0110847-g002\" target=\"_blank\">Figure 2</a> was 60.2±17 nM.</p><p>Kinetic parameters extracted from fitting the binding curves of concentration series of three different mAbs against a single preparation of immobilized CXCR4 ACMs.</p>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209990, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0110847.t001", "stats"=>{"downloads"=>1, "page_views"=>7, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Kinetic_parameters_extracted_from_fitting_the_binding_curves_of_concentration_series_of_three_different_mAbs_against_a_single_preparation_of_immobilized_CXCR4_ACMs_/1209990", "title"=>"Kinetic parameters extracted from fitting the binding curves of concentration series of three different mAbs against a single preparation of immobilized CXCR4 ACMs.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2014-10-20 02:48:45"}
  • {"files"=>["https://ndownloader.figshare.com/files/1724379"], "description"=>"<p>The analyte was injected in triplicate, at cycle 7, 14, and 21. Intermediate cycles involved blank injections. The blue line shows the fit to the curve assuming 1∶1 binding kinetics. The inset shows the relative decrease in binding activity of the surface as measured by the binding level 4 s before the end of the injection.</p>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209987, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0110847.g003", "stats"=>{"downloads"=>1, "page_views"=>26, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Sensorgrams_of_mAb_12G5_binding_100_nM_to_CXCR4_ACMs_immobilized_at_low_RU_ca_1400_via_biotin_spteptavidin_immobilization_of_the_embedding_polymersome_matrix_/1209987", "title"=>"Sensorgrams of mAb 12G5 binding (100 nM) to CXCR4-ACMs immobilized at low RU (ca. 1400) via biotin/spteptavidin immobilization of the embedding polymersome matrix.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-10-20 02:48:45"}
  • {"files"=>["https://ndownloader.figshare.com/files/1724381"], "description"=>"<p>Conditions were the same as in <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110847#pone-0110847-g002\" target=\"_blank\">Figure 2</a>, except for the immobilization level, which is as indicated in the figure.</p>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209989, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0110847.g004", "stats"=>{"downloads"=>2, "page_views"=>28, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_SPR_sensorgrams_obtained_by_kinetic_screening_of_three_different_types_of_conformationally_sensitive_ABs_to_a_single_CXCR4_ACM_preparation_immobilized_on_a_sensor_chip_/1209989", "title"=>"SPR sensorgrams obtained by kinetic screening of three different types of conformationally sensitive ABs to a single CXCR4 ACM preparation immobilized on a sensor chip.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-10-20 02:48:45"}
  • {"files"=>["https://ndownloader.figshare.com/files/1724375"], "description"=>"<p>A: PB-PEO polymersomes and CXCR4 c-DNA were subjected to in-vitro synthesis using a wheat germ coupled translation-transcription extract (WGE) and purified by a filtration step. After filtration, insertion of CXCR4 in the polymersome membrane was verified by Western blot. CXCR4 c-DNA in absence of polymersomes, which went through the same process, did not show the presence of CXCR4. The positive control is a commercially available CXCR4 cell membrane preparation. B: After purification, ACMs were immobilized onto a biosensor gold chip by first coupling streptavidin using standard EDC/NHS coupling, and then capturing the CXCR4 ACMs by the interaction with streptavidin of biotinylated lipid mixed into the polymersome membrane.</p>", "links"=>[], "tags"=>["binding behaviour", "drug targets", "cxcr", "Kinetic characterization", "receptor", "gpcr", "biosensor chips", "diblock copolymer membranes", "expression system", "Conformational Antibody Binding"], "article_id"=>1209983, "categories"=>["Biological Sciences"], "users"=>["Hans-Peter M. de Hoog", "Esther M. Lin JieRong", "Sourabh Banerjee", "Fabien M. Décaillot", "Madhavan Nallani"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0110847.g001", "stats"=>{"downloads"=>0, "page_views"=>14, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_In_vitro_synthesis_and_direct_insertion_of_CXCR4_into_polymersomes_/1209983", "title"=>"In-vitro synthesis and direct insertion of CXCR4 into polymersomes.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-10-20 02:48:45"}

PMC Usage Stats | Further Information

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Relative Metric

{"start_date"=>"2014-01-01T00:00:00Z", "end_date"=>"2014-12-31T00:00:00Z", "subject_areas"=>[{"subject_area"=>"/Biology and life sciences/Neuroscience", "average_usage"=>[289]}, {"subject_area"=>"/Engineering and technology", "average_usage"=>[282]}, {"subject_area"=>"/Engineering and technology/Equipment", "average_usage"=>[287, 444]}, {"subject_area"=>"/Physical sciences", "average_usage"=>[271]}, {"subject_area"=>"/Social sciences", "average_usage"=>[318]}, {"subject_area"=>"/Social sciences/Psychology", "average_usage"=>[314, 476]}]}
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