Transduction of Glycan-Lectin Binding Using Near-Infrared Fluorescent Single-Walled Carbon Nanotubes for Glycan Profiling

There is significant interest in developing new detection platforms for characterizing glycosylated proteins, despite the lack of easily synthesized model glycans or high affinity receptors for this anal. problem. In this work, the authors demonstrate a sensor array employing recombinant lectins as glycan recognition sites tethered via Histidine tags to Ni2+ complexes that act as fluorescent quenchers for semiconducting single-walled carbon nanotubes (SWNTs) embedded in a chitosan hydrogel spot to measure binding kinetics of model glycans. The authors examine, as model glycans, both free and streptavidin-tethered biotinylated monosaccharides. Two higher-affined glycan-lectin pairs are explored: fucose (Fuc) to PA-IIL and N-acetylglucosamine (GlcNAc) to GafD. The dissocn. consts. (KD) for these pairs as free glycans (106 and 19 μM, resp.) and streptavidin-tethered (142 and 50 μM resp.) were found. The abs. detection limit for the current platform was 2 μg of glycosylated protein or 100 ng of free glycan to 20 μg of lectin. Glycan detection (GlcNAc-streptavidin at 10 μM) is demonstrated at the single nanotube level as well by monitoring the fluorescence from individual SWNT sensors tethered to GafD lectin. Over a population of 1000 nanotubes, 289 of the SWNT sensors had signals strong enough to yield kinetic information (KD of 250±10 μM). The authors are also able to identify the locations of "strong transducers" on the basis of dissocn. const. (four sensors with KD < 10 μM) or overall signal modulation (eight sensors with >5% quench response). The authors report the key finding that the brightest SWNTs are not the best transducers of glycan binding. SWNTs ranging in intensity between 50 and 75% of the max. show the greatest response. The ability to pinpoint strong-binding, single sensors is promising to build a nanoarray of glycan-lectin transducers as a high throughput method to profile glycans without protein labeling or glycan liberation pretreatment steps. [on SciFinder(R)]

Published in:
Journal of the American Chemical Society, 133, 17923-17933

 Record created 2015-03-03, last modified 2018-12-03

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