Details zur Publikation

Kategorie Textpublikation
Referenztyp Zeitschriften
DOI 10.1002/cbic.201100774
Titel (primär) Kinetic and stoichiometric characterisation of streptavidin-binding aptamers
Autor Ruigrok, V.J.B.; van Duijn, E.; Barendregt, A.; Dyer, K.; Tainer, J.A.; Stoltenburg, R.; Strehlitz, B.; Levisson, M.; Smidt, H.; van der Oost, J.
Quelle Chembiochem
Erscheinungsjahr 2012
Department UBZ
Band/Volume 13
Heft 6
Seite von 829
Seite bis 836
Sprache englisch
Keywords aptamers; native mass spectrometry; SAXS; SELEX; surface plasmon resonance

Aptamers are oligonucleotide ligands that are selected for high-affinity binding to molecular targets. Only limited knowledge relating to relations between structural and kinetic properties that define aptamer–target interactions is available. To this end, streptavidin-binding aptamers were isolated and characterised by distinct analytical techniques. Binding kinetics of five broadly similar aptamers were determined by surface plasmon resonance (SPR); affinities ranged from 35–375 nM with large differences in association and dissociation rates. Native mass spectrometry showed that streptavidin can accommodate up to two aptamer units. In a 3D model of one aptamer, conserved regions are exposed, strongly suggesting that they directly interact with the biotin-binding pockets of streptavidin. Mutational studies confirmed both conserved regions to be crucial for binding. An important result is the observation that the most abundant aptamer in our selections is not the tightest binder, emphasising the importance of having insight into the kinetics of complex formation. To find the tightest binder it might be better to perform fewer selection rounds and to focus on post-selection characterisation, through the use of complementary approaches as described in this study.
dauerhafte UFZ-Verlinkung
Ruigrok, V.J.B., van Duijn, E., Barendregt, A., Dyer, K., Tainer, J.A., Stoltenburg, R., Strehlitz, B., Levisson, M., Smidt, H., van der Oost, J. (2012):
Kinetic and stoichiometric characterisation of streptavidin-binding aptamers
ChemBioChem 13 (6), 829 - 836 10.1002/cbic.201100774