Site-directed conjugation of single-stranded DNA to affinity proteins: quantifying the importance of conjugation strategy
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Date
2024-06-21
Publication Type
Journal Article
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yes
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Abstract
Affinity protein-oligonucleotide conjugates are increasingly being explored as diagnostic and therapeutic tools. Despite growing interest, these probes are typically constructed using outdated, non-selective chemistries, and little has been done to investigate how conjugation to oligonucleotides influences the function of affinity proteins. Herein, we report a novel site-selective conjugation method for furnishing affinity protein-oligonucleotide conjugates in a 93% yield within fifteen minutes. Using SPR, we explore how the choice of affinity protein, conjugation strategy, and DNA length impact target binding and reveal the deleterious effects of non-specific conjugation methods. Furthermore, we show that these adverse effects can be minimised by employing our site-selective conjugation strategy, leading to improved performance in an immuno-PCR assay. Finally, we investigate the interactions between affinity protein-oligonucleotide conjugates and live cells, demonstrating the benefits of site-selective conjugation. This work provides critical insight into the importance of conjugation strategy when constructing affinity protein-oligonucleotide conjugates.
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published
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Journal / series
Volume
15 (23)
Pages / Article No.
8982 - 8992
Publisher
Royal Society of Chemistry
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Organisational unit
03914 - deMello, Andrew / deMello, Andrew
09736 - Aceto, Nicola / Aceto, Nicola
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Funding
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