## 3 Results and Discussion
### Binding Interaction Analysis with PLIP
#### 3.5.1 Interactions with Estrogen Receptor
For CGAMut4, eighteen hydrogen bonds and twelve hydrophobic interactions were observed, while two salt bridges were formed between His377A (Supplementary Information Table S3q and Figure 3q). New H-bonds formed include those with Thr371B, Asp374B, Arg412C, and Ser536B. New hydrophobic interactions include those with Val422C, Lys531B, Asp538B, Leu541A, and Glu542A. No \(\pi\)-stacking or \(\pi\)-cation interactions were formed. The CSAMut4 conjugate (Supplementary Information Table S3r and Figure 3r) showed fourteen H-bonds and nine hydrophobic interactions. Nearly all H-bonds observed were formed with different residues compared to those for Mut4 alone; interacting residues include Arg412D, Glu423D, His516D, Lys520D, and Ser536A. New hydrophobic interactions included those with Thr460B, Leu462B, and Leu539A. \(\pi\)-stacking and salt bridges seen for neat Mut4 were not maintained. A \(\pi\)-cation interaction was observed with Arg412D. For GLAMut4, the conjugate exhibited nineteen H-bonds and eleven hydrophobic interactions (Supplementary Information Table S3s and Figure 3s). New H-bonds were seen with Arg412D, Asp426C, Met427D, Leu462B, Ser463B, His516C, Lys520C, Ser536A, and Glu542A, while new hydrophobic interactions occurred with Glu380A, Val422D, and Asp538A. A \(\pi\)-stacking interaction was formed with Tyr537B and a \(\pi\)-cation interaction was seen with His516C. For the RMAMut4 conjugate, additional H-bonding (twenty-two total) was observed and slightly reduced hydrophobic interactions (nine) occurred compared to Mut4 alone (Supplementary Information Table S3t and Figure 3t). New H-bonds included those with Cys381B, Trp383B, Arg412D, Asp426D, His516D, Ser518B, and Asn519B and new hydrophobic interactions included those with Glu380B, Thr460B, and Ser463B. \(\pi\)-stacking interactions were formed with His516D and Tyr537A, a \(\pi\)-cation interaction was observed with His377B, and salt bridges occurred with Arg412D and Lys520D. These results indicated that all of the polyphenol-peptide conjugates, as well as the peptides, interacted with the ER-\(\alpha\); furthermore, the conjugation of select peptides with some of the polyphenols enhanced binding. Specifically, the binding interactions of Mut1 was improved by conjugation with gallate, while that of Mut2 improved upon conjugation with all four of the polyphenols. In the case of Mut3, improved binding interactions were observed with CGA, while, for Mut4, conjugation with gallate enhanced interactions. The relatively stronger binding interactions between chlorogeneate conjugates and ER\(\alpha\) were likely resultant of the hydrogen bonding ability of CGA. In comparison to CGA, carnosate is relatively hydrophobic due to its extensive ring system, which likely contributed to the weaker binding interactions observed for carnosate conjugates with ER-\(\alpha\). It is interesting to note that, among the mutated peptides, Mut3 and Pep had very similar binding interactions, indicating that changing Y-H-W-Y-G-Y-T-P-Q-N to Y-H-W-Y-G-Y-T-H-Q-N did not have a large impact on binding, though His is significantly different in properties from proline. This can be attributed to the fact that both peptides showed an equal number of H-bonding interactions with the ER-\(\alpha\), indicating that H-bonding played a key role in overall binding interactions.