Version 1
: Received: 23 September 2024 / Approved: 24 September 2024 / Online: 24 September 2024 (14:04:05 CEST)
How to cite:
Banerjee, S.; Fraser, K.; Crone, D. E.; Patel, J. C.; Bondos, S.; Bystroff, C. Challenges and Solutions for Leave-One-Out Biosensor Design in the Context of a Rugged Fitness Landscape. Preprints2024, 2024091923. https://doi.org/10.20944/preprints202409.1923.v1
Banerjee, S.; Fraser, K.; Crone, D. E.; Patel, J. C.; Bondos, S.; Bystroff, C. Challenges and Solutions for Leave-One-Out Biosensor Design in the Context of a Rugged Fitness Landscape. Preprints 2024, 2024091923. https://doi.org/10.20944/preprints202409.1923.v1
Banerjee, S.; Fraser, K.; Crone, D. E.; Patel, J. C.; Bondos, S.; Bystroff, C. Challenges and Solutions for Leave-One-Out Biosensor Design in the Context of a Rugged Fitness Landscape. Preprints2024, 2024091923. https://doi.org/10.20944/preprints202409.1923.v1
APA Style
Banerjee, S., Fraser, K., Crone, D. E., Patel, J. C., Bondos, S., & Bystroff, C. (2024). Challenges and Solutions for Leave-One-Out Biosensor Design in the Context of a Rugged Fitness Landscape. Preprints. https://doi.org/10.20944/preprints202409.1923.v1
Chicago/Turabian Style
Banerjee, S., Sarah Bondos and Christopher Bystroff. 2024 "Challenges and Solutions for Leave-One-Out Biosensor Design in the Context of a Rugged Fitness Landscape" Preprints. https://doi.org/10.20944/preprints202409.1923.v1
Abstract
The Leave-One-Out (LOO) Green Fluorescent Protein (GFP) approach to biosensor design com-bines computational protein design with split protein reconstitution. LOO-GFPs reversibly fold and gain fluorescence upon encountering its target peptide, which can be redefined by computa-tional design of the LOO site. Such an approach can be used to create reusable biosensors for the early detection of emerging biological threats. Enlightening biophysical inferences for nine LOO-GFP biosensor libraries are presented, with target sequences from dengue, influenza, or HIV, replacing beta strands 7, 8 or 11. An initially low hit rate was traced to components of the energy function, manifesting in the over-rewarding of over-tight side chain packing. Also, screening by colony picking required a low library complexity, but designing a biosensor against a peptide of at least 12 residues requires a high complexity library. This double-bind was solved using a "piecemeal" iterative design strategy. Also, designed LOO-GFPs fluoresced in the unbound state due to unwanted dimerization, but this was solved by fusing a fully functional prototype LOO-GFP to a fiber-forming protein, Drosophila ultrabithorax, creating a biosensor fiber. One in-fluenza hemagglutinin biosensor is characterized here in detail, showing a shifted excita-tion/emission spectrum, a micromolar affinity for the target peptide, and an unexpected pho-to-switching ability.
Keywords
green fluorescent protein; biosensors; folding; library screening; biosensor materials; computational design; folding energy landscape; chromophore; ultrabithorax fibers
Subject
Biology and Life Sciences, Biophysics
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
