Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography
The paradigm of drug discovery have been to find the strongest possible binder to the target by high-throughput screening (HTS) but high affinity interactions are related to low kinetic off rates and thus result in severe side-effects and non-approved drugs. Lead molecules working in a transient man...
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Linnéuniversitetet, Institutionen för naturvetenskap, NV
2012
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ndltd-UPSALLA1-oai-DiVA.org-lnu-199742013-01-08T13:41:18ZEvaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity ChromatographyengJönsson, DanielLinnéuniversitetet, Institutionen för naturvetenskap, NV2012peak profile methoddissociation / association rate constantsweak affinity chromatographyband-broadeningChemistryKemiThe paradigm of drug discovery have been to find the strongest possible binder to the target by high-throughput screening (HTS) but high affinity interactions are related to low kinetic off rates and thus result in severe side-effects and non-approved drugs. Lead molecules working in a transient manner (KD > µM) will allow for rapid off rates and possibly less side-effects. In this study the peak profile method applied to weak affinity chromatography (WAC) was evaluated as a simple way to provide the kinetics of the interaction and thereby allowing for high-throughput determinations. In the peak profile formula all band-broadening effects except the stationary mass transfer is subtracted which simplifies the calculations for the kinetics of the interaction tremendously. The technique was evaluated by screening of 3 different benzamidines at 3 linear flow-rates using zonal chromatography and human α-thrombin as immobilized target protein. The kinetics of the interaction could unfortunately not be determined. This was possibly due to the flow-rates not being high enough as indicated by a low critical ratio (η < 1). Higher flow-rates would increase the contribution to band-broadening due to kinetic effects but would also require more precise estimation of peak variance. Student thesisinfo:eu-repo/semantics/bachelorThesistexthttp://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-19974application/pdfinfo:eu-repo/semantics/openAccess |
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peak profile method dissociation / association rate constants weak affinity chromatography band-broadening Chemistry Kemi |
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peak profile method dissociation / association rate constants weak affinity chromatography band-broadening Chemistry Kemi Jönsson, Daniel Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
description |
The paradigm of drug discovery have been to find the strongest possible binder to the target by high-throughput screening (HTS) but high affinity interactions are related to low kinetic off rates and thus result in severe side-effects and non-approved drugs. Lead molecules working in a transient manner (KD > µM) will allow for rapid off rates and possibly less side-effects. In this study the peak profile method applied to weak affinity chromatography (WAC) was evaluated as a simple way to provide the kinetics of the interaction and thereby allowing for high-throughput determinations. In the peak profile formula all band-broadening effects except the stationary mass transfer is subtracted which simplifies the calculations for the kinetics of the interaction tremendously. The technique was evaluated by screening of 3 different benzamidines at 3 linear flow-rates using zonal chromatography and human α-thrombin as immobilized target protein. The kinetics of the interaction could unfortunately not be determined. This was possibly due to the flow-rates not being high enough as indicated by a low critical ratio (η < 1). Higher flow-rates would increase the contribution to band-broadening due to kinetic effects but would also require more precise estimation of peak variance. |
author |
Jönsson, Daniel |
author_facet |
Jönsson, Daniel |
author_sort |
Jönsson, Daniel |
title |
Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
title_short |
Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
title_full |
Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
title_fullStr |
Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
title_full_unstemmed |
Evaluation of Rate Constants from Protein-Ligand Interactions with Weak Affinity Chromatography |
title_sort |
evaluation of rate constants from protein-ligand interactions with weak affinity chromatography |
publisher |
Linnéuniversitetet, Institutionen för naturvetenskap, NV |
publishDate |
2012 |
url |
http://urn.kb.se/resolve?urn=urn:nbn:se:lnu:diva-19974 |
work_keys_str_mv |
AT jonssondaniel evaluationofrateconstantsfromproteinligandinteractionswithweakaffinitychromatography |
_version_ |
1716526086348603392 |