**5. Conclusions**

With the development of machine learning in biomolecular modeling and characterization, knowledge of interaction-dependent features in spectral data of SERS-measured molecules and structures leads to dynamic single-molecule SERS fingerprints. We have shown that the dynamic vibrational motion analysis of a single molecule can investigate the Van der Waals interaction.

The findings presented in the current research prove the MD simulation's applicability for transient vibrational spectra of biomolecular building blocks such as nucleotides or amino acids. We have shown that the obtained vibrational spectra are sensitive enough to reflect even weak Van der Waals interactions in the few component systems studied, such as nucleotide–Au-NP–graphene. The transient regime of nucleotide passing by NP and through the graphene was shown to be spectrally sensitive. This is confirmed by changes in the bands of amino and methyl groups attached to the rings dependent on interaction strength and length. The transient vibrational spectra record enables discriminating different interaction events with the spectral fingerprints of molecules and NP that also exhibit spectral modification. We consider expanding the scope of the method for protein fragments and conformations. The use of interaction-dependent MD and ab initio

MD simulations of transient spectra can make the measurement of SERS of unattached molecules attainable in a small number of pulses.

**Supplementary Materials:** The following are available online at https://www.mdpi.com/article/10 .3390/bios11020037/s1. Table S1: Frequencies of cytosine nucleotide bonds, highest intensities. Table S2: Frequencies of thymine nucleotide bonds, highest intensities. Table S3: Frequencies of adenine nucleotide bonds, highest intensities. Table S4: Frequencies of guanine nucleotide bonds, highest intensities. Figure S1: The numbering of the nucleotide atoms used in Tables S1–S4 and S5–S8. Table S5: Frequencies of cytosine base bonds, highest intensities. Table S6: Frequencies of thymine base bonds, highest intensities. Table S7: Frequencies of adenine base bonds, highest intensities. Table S8: Frequencies of guanine base bonds, highest intensities.

**Author Contributions:** Conceptualization, methodology, software development, validation, supervision: T.Z.; data curation: M.Y. and S.I.; writing—original draft preparation, review, and editing: T.Z. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** Not applicable.

**Acknowledgments:** T.Z. gratefully acknowledges the technical assistance in the nucleotide calculations and preparation of the results of Appendix A done by H. Mizuguchi and T. Kitani. All calculations were performed at Applied Mechano-Informatics Laboratory in Toyama University.

**Conflicts of Interest:** The authors declare no conflict of interest.
