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Click-Particle Display for Base-Modified Aptamer Discovery
journal contribution
posted on 2019-10-12, 12:13 authored by Chelsea
K. L. Gordon, Diana Wu, Anusha Pusuluri, Trevor A. Feagin, Andrew T. Csordas, Michael S. Eisenstein, Craig J. Hawker, Jia Niu, Hyongsok Tom SohBase-modified aptamers
that incorporate non-natural chemical moieties
can achieve greatly improved affinity and specificity relative to
natural DNA or RNA aptamers. However, conventional methods for generating
base-modified aptamers require considerable expertise and resources.
In this work, we have accelerated and generalized the process of generating
base-modified aptamers by combining a click-chemistry strategy with
a fluorescence-activated cell sorting (FACS)-based screening methodology
that measures the affinity and specificity of individual aptamers
at a throughput of ∼107 per hour. Our “click-particle
display (PD)” strategy offers many advantages. First, almost
any chemical modification can be introduced with a commercially available
polymerase. Second, click-PD can screen vast numbers of individual
aptamers on the basis of quantitative on- and off-target binding measurements
to simultaneously achieve high affinity and specificity. Finally,
the increasing availability of FACS instrumentation in academia and
industry allows for easy adoption of click-PD in a broader scientific
community. Using click-PD, we generated a boronic acid-modified aptamer
with ∼1 μM affinity for epinephrine, a target for which
no aptamer has been reported to date. We subsequently generated a
mannose-modified aptamer with nanomolar affinity for the lectin concanavalin
A (Con A). The strong affinity of both aptamers is fundamentally dependent
upon the presence of chemical modifications, and we show that their
removal essentially eliminates aptamer binding. Importantly, our Con
A aptamer exhibited exceptional specificity, with minimal binding
to other structurally similar lectins. Finally, we show that our aptamer
has remarkable biological activity. Indeed, this aptamer is the most
potent inhibitor of Con A-mediated hemagglutination reported to date.