Comparative analyses of a small molecule/enzyme interaction by multiple users of Biacore technology
| العنوان: | Comparative analyses of a small molecule/enzyme interaction by multiple users of Biacore technology |
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| المؤلفون: | Stefano F. Liparoto, Melanie Wong, Eric S. Day, Michael Sekar, Donald B. Bennett, Michelle J. Cannon, Dave Casper, Edward J. Collins, Susanne Nyholm Westin, Inna Gorshkova, Yue Jin Li, Karen M. Worthy, Amy Neurauter, Daulet Satpaev, Gerardo R. Marchesini, Paul J. Cachia, Teresa Chu, Ryan James Darling, Jack Simpson, Daniel Kao, Dax A. Rice, Karlicek Shannon Marie, Robert J. Fisher, Magdalena Bynum, Michael Swanson, David G. Myszka, Eric D. Sullivan, Dianne Hodges, Lindsey R. Roberts, Amy M. Liang, Ashique Rafique, Huawei Qiu, Andrew G. Stephen, Giuseppe A. Papalia, Iva Navratilova, Eric Hommema, Béatrice Luginbühl, Jean Inman, Stephen J. Ullrich |
| المصدر: | Analytical Biochemistry 330 (2004) 1 Analytical Biochemistry, 330(1), 98-113 |
| بيانات النشر: | Elsevier BV, 2004. |
| سنة النشر: | 2004 |
| مصطلحات موضوعية: | Analyte, binding, mass-transport, Immobilized enzyme, RIKILT - Business Unit Veiligheid & Gezondheid, Biophysics, Analytical chemistry, Carbonic Anhydrase II, Biochemistry, Chemical kinetics, Reaction rate constant, surface, Molecular Biology, Observer Variation, Chromatography, plasmon resonance biosensors, Chemistry, carbonic-anhydrase, Enzyme Interaction, Interaction model, Cell Biology, Surface Plasmon Resonance, kinetic-analysis, Small molecule, Research Personnel, rate constants, Acetazolamide, Dissociation constant, Kinetics, RIKILT - Business Unit Safety & Health, Protein Binding |
| الوصف: | To gauge the experimental variability associated with Biacore analysis, 36 different investigators analyzed a small molecule/enzyme interaction under similar conditions. Acetazolamide (222 g/mol) binding to carbonic anhydrase II (CAII; 30,000 Da) was chosen as a model system. Both reagents were stable and their interaction posed a challenge to measure because of the low molecular weight of the analyte and the fast association rate constant. Each investigator created three different density surfaces of CAII and analyzed an identical dilution series of acetazolamide (ranging from 4.1 to 1000 nM). The greatest variability in the results was observed during the enzyme immobilization step since each investigator provided their own surface activating reagents. Variability in the quality of the acetazolamide binding responses was likely a product of how well the investigators’ instruments had been maintained. To determine the reaction kinetics, the responses from the different density surfaces were fit globally to a 1:1 interaction model that included a term for mass transport. The averaged association and dissociation rate constants were 3.1 ± 1.6 × 10 6 M −1 s −1 and 6.7 ± 2.5 × 10 −2 s −1 , respectively, which corresponded to an average equilibrium dissociation constant ( K D ) of 2.6 ± 1.4 × 10 −8 M. The results provide a benchmark of variability in interpreting binding constants from the biosensor and highlight keys areas that should be considered when analyzing small molecule interactions. |
| وصف الملف: | application/pdf; application/octet-stream |
| تدمد: | 0003-2697 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::2f902d2fc2f47b9116e87905fdebf021 https://doi.org/10.1016/j.ab.2004.02.027 |
| حقوق: | RESTRICTED |
| رقم الأكسشن: | edsair.doi.dedup.....2f902d2fc2f47b9116e87905fdebf021 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 00032697 |
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