Streptavidin-Biotin Binding is Intrinsically Non-Cooperative, but Experimental Artefacts Can Induce Apparent Positive Cooperativity
Abstract
The biotin-streptavidin interaction is a key component of nanobiotechnology, yet the nature of its cooperativity remains a debated question. Here, we introduce a label-free spectroscopy approach based on confocal UV autofluorescence microscopy to probe the binding cooperativity of this high-affinity system. By leveraging the intrinsic autofluorescence of tryptophan residues, we avoid artefacts associated with extrinsic fluorescent labels, enabling precise measurements of fully biotin-saturated streptavidin across a wide range of molar ratios. Our experimental results, supported by Monte Carlo simulations, provide conclusive evidence for the absence of cooperativity. However, we show that false signs of apparent positive cooperativity can emerge from local concentration inhomogeneities occurring when rapid reaction kinetics outpace mixing. By establishing the non-cooperative nature of biotin-streptavidin binding and identifying the key conditions to avoid cooperativity artefacts, these insights resolve a longstanding debate and offer critical implications for streptavidin-based biotechnological applications.
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BibTeX
@misc{veedu2026streptavidinbiotin,
title = {{Streptavidin-Biotin Binding is Intrinsically Non-Cooperative, but Experimental Artefacts Can Induce Apparent Positive Cooperativity}},
author = {Malavika Kayyil Veedu and Eléonore Martenne Duplan and Saurabh Rai and Prithu Roy and Jérôme Wenger},
howpublished = {arXiv},
year = {2026},
month = oct,
doi = {10.48550/arxiv.2610.41363},
eprint = {2610.41363},
archivePrefix = {arXiv},
url = {https://arxiv.org/abs/2610.41363},
note = {CC BY 4.0},
}
RIS
TY - UNPB
TI - Streptavidin-Biotin Binding is Intrinsically Non-Cooperative, but Experimental Artefacts Can Induce Apparent Positive Cooperativity
AU - Malavika Kayyil Veedu
AU - Eléonore Martenne Duplan
AU - Saurabh Rai
AU - Prithu Roy
AU - Jérôme Wenger
PY - 2026
DA - 2026/10/30
PB - arXiv
DO - 10.48550/arxiv.2610.41363
UR - https://arxiv.org/abs/2610.41363
AB - The biotin-streptavidin interaction is a key component of nanobiotechnology, yet the nature of its cooperativity remains a debated question. Here, we introduce a label-free spectroscopy approach based on confocal UV autofluorescence microscopy to probe the binding cooperativity of this high-affinity system. By leveraging the intrinsic autofluorescence of tryptophan residues, we avoid artefacts associated with extrinsic fluorescent labels, enabling precise measurements of fully biotin-saturated streptavidin across a wide range of molar ratios. Our experimental results, supported by Monte Carlo simulations, provide conclusive evidence for the absence of cooperativity. However, we show that false signs of apparent positive cooperativity can emerge from local concentration inhomogeneities occurring when rapid reaction kinetics outpace mixing. By establishing the non-cooperative nature of biotin-streptavidin binding and identifying the key conditions to avoid cooperativity artefacts, these insights resolve a longstanding debate and offer critical implications for streptavidin-based biotechnological applications.
ER -