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We address the challenge of understanding how hydrophobic interactions are encoded by fusion peptide sequences within coronavirus (CoV) spike proteins. Within the fusion peptides of SARS-CoV-2 and MERS-CoV, a largely conserved peptide sequence called FP1 (SFIEDLLFNK and SAIEDLLFDK in SARS-2 and MERS, respectively) has been proposed to play a key role in encoding hydrophobic interactions that drive viral-host cell membrane fusion. While a non-polar triad (LLF) is common to both FP1 sequences, and thought to dominate the encoding of hydrophobic interactions, FP1 from SARS and MERS differ in two residues (Phe 2 versus Ala 2 and Asn 9 versus Asp 9s, respectively). Here we explore if single molecule force measurements can quantify hydrophobic interactions encoded by FP1 sequences, and then ask if sequence variations between FP1 from SARS-2 and MERS lead to significant differences in hydrophobic interactions. We find that both SARS-2 and MERS wild-type FP1 generate measurable hydrophobic interactions at the single molecule level, but that SARS-2 FP1 encodes a substantially stronger hydrophobic interaction than its MERS counterpart (1.91 ± 0.03 nN versus 0.68 ± 0.03 nN, respectively). By performing force measurements with FP1 sequences with single amino acid substitutions, we determine that a single residue mutation (Phe 2 versus Ala 2) causes the almost threefold difference in the hydrophobic interaction strength generated by the FP1 of SARS-2 versus MERS, despite the presence of LLF in both sequences. Infrared spectroscopy and circular dichroism measurements support the proposal that the outsized influence of Phe 2 versus Ala 2 on the hydrophobic interaction arises from variation in the secondary structure adopted by FP1. Overall, these insights reveal how single residue diversity in viral fusion peptides, including FP1 of SARS-CoV-2 and MERS-CoV, can lead to substantial changes in intermolecular interactions proposed to play a key role in viral fusion, and hint at strategies for regulating hydrophobic interactions of peptides in a range of contexts. 

Hydrophobic interactions play a central role in bioinspired strategies for molecular self-assembly in water, yet how these interactions are encoded by chemically heterogeneous interfaces is poorly understood. We report an experimental investigation of the influence of immobilized polar groups (amine) and cations (ammonium and guanidinium) on enthalpic and entropic contributions to hydrophobic interactions mediated by methyl-terminated surfaces at temperatures ranging from 298 K to 328 K and pH values between 3.5 to 10.5. We use our measurements to calculate the change in free energy (and enthalpic and entropic components) that accompanies transfer of each surface from aqueous TEA containing 60 vol% methanol into aqueous TEA ( i.e. , transfer free energy that characterizes hydrophobicity). We find the thermodynamic signature of the pure methyl surface (positive transfer enthalpy and entropy) to be altered qualitatively by incorporation of amine or guanidinium groups into the surface (negative transfer enthalpy and near zero transfer entropy). In contrast, ammonium groups immobilized on a methyl surface do not change the thermodynamic signature of the hydrophobic interaction. Compensation of entropy and enthalpy is clearly evident in our results, but the overall trends in the transfer free energies are dominated by enthalpic effects. This observation and others lead us to hypothesize that the dominant effect of the immobilized charged or polar groups in our experiments is to influence the number or strength of hydrogen bonds formed by interfacial water molecules adjacent to the nonpolar domains. Overall, these results provide insight into entropy–enthalpy compensation at chemically heterogeneous surfaces, and generate hypotheses and a rich experimental dataset for further exploration via simulation.