Drug Discovery at the Single Molecule Level: Inhibition-in-Solution Assay of Membrane-Reconstituted beta-Secretase Using Single-Molecule Imaging
Artikel i vetenskaplig tidskrift, 2015

Inhibition-in-solution assays (ISA) employing surface-based biosensors such as surface plasmon resonance (SPR) are an effective screening approach in drug discovery. However, analysis of potent binders remains a significant hurdle due to limited sensitivity and accompanied depletion of the inhibiting compounds due to high protein concentrations needed for detectable binding signals. To overcome this limitation, we explored a microscopy-based single-molecule ISA compatible with liposome-reconstituted membrane proteins. Using a set of validated small molecule inhibitors against beta-secretase 1 (BACE1), the assay was benchmarked with respect to sensitivity and dynamic range against SPR. We demonstrate that the dynamic range of measurable affinities is greatly extended by more than 2 orders of magnitude as compared to SPR, thus facilitating measurements of highly potent (K-d < nM) compounds.


Anders Gunnarsson

AstraZeneca Sweden

Arjan Snijder

AstraZeneca Sweden

J. Hicks

AstraZeneca Sweden

J. Gunnarsson

AstraZeneca Sweden

Fredrik Höök

Chalmers, Teknisk fysik, Biologisk fysik

S. Geschwindner

AstraZeneca Sweden

Analytical Chemistry

0003-2700 (ISSN) 1520-6882 (eISSN)

Vol. 87 4100-4103


Analytisk kemi