Brookes, Jennifer Clare;
Gray, Eleanor Ruth;
Loynachan, Colleen Noel;
Gut, Michelle Jennifer;
Miller, Benjamin Saul;
Stanislas Brogan, Alex Paul;
McKendry, Rachel Anne;
(2022)
Thermodynamic analysis of an entropically driven, high affinity nanobody-HIV p24 interaction.
Biophysical Journal
10.1016/j.bpj.2022.12.019.
(In press).
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Abstract
Protein-protein interactions are fundamental to life processes. Complementary computational, structural and biophysical studies of these interactions enable the forces behind their specificity and strength to be understood. Antibody fragments such as single-chain antibodies have the specificity and affinity of full antibodies but a fraction of their size, expediting whole molecule studies and distal effects without exceeding the computational capacity of modelling systems. We previously reported the crystal structure of a high affinity nanobody 59H10 bound to HIV-1 capsid protein p24, and deduced key interactions using all-atom molecular dynamics (MD) simulations. We studied the properties of closely-related medium (37E7) and low (48G11) affinity nanobodies, to understand how changes of three (37E7) or one (48G11) amino acids impacted these interactions, however, the contributions of enthalpy and entropy were not quantified. Here, we report the use of qualitative and quantitative experimental and in silico approaches to separate the contributions of enthalpy and entropy. We used complementary circular dichroism spectroscopy and molecular dynamics simulations to qualitatively delineate changes between nanobodies in isolation and complexed with p24. Using quantitative techniques such as isothermal titration calorimetry alongside WaterMap and Free Energy Perturbation protocols, we found the difference between high (59H10) and medium (37E7) affinity nanobodies on binding to HIV-1 p24 is entropically driven, accounted for by the release of unstable waters from the hydrophobic surface of 59H10. Our results provide an exemplar of the utility of parallel in vitro and in silico studies and highlight that differences in entropic interactions between amino acids and water molecules are sufficient to drive orders of magnitude differences in affinity.
| Type: | Article |
|---|---|
| Title: | Thermodynamic analysis of an entropically driven, high affinity nanobody-HIV p24 interaction |
| Location: | United States |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1016/j.bpj.2022.12.019 |
| Publisher version: | https://doi.org/10.1016/j.bpj.2022.12.019 |
| Language: | English |
| Additional information: | © 2022 Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/ |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices > UCL BEAMS UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences > London Centre for Nanotechnology |
| URI: | https://discovery-pp.ucl.ac.uk/id/eprint/10162582 |
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