Metadynamics and Raman Spectroscopy for Glycan Structure-Spectrum Mapping

Raman spectroscopy offers label-free access to molecular vibrations across biological systems, from small molecules to flexible biomolecular oligomers and whole cells. Yet, understanding how conformational ensembles encode vibrational spectra remains challenging. While small-molecule Raman spectra often permit direct structural interpretation, intermediate-sized biomolecules such as glycans, short nucleic acids, and peptides exhibit compositional and conformational heterogeneity that obscures structure-spectrum relationships. We introduce DynaSpec, an ensemble-resolved structure-spectrum mapping framework that links equilibrium conformational populations to vibrational observables. DynaSpec integrates metadynamics in a machine-learned latent space, population-weighted DFT Raman calculations, and experimental spectroscopy to interpret congested Raman signatures mechanistically. By connecting normal modes to the internal structural coordinates that modulate them, DynaSpec enables bidirectional structure-spectrum inference. Using glycans as a stringent test case, ensemble-averaged DFT predictions show strong agreement with experimental Raman spectra and support >85% classification accuracy across 13 N-glycans. The analysis reveals a conserved α1,6 open-curled switching motif across N-glycans and identifies a glycosidic torsion-enriched spectral window that enables experimental discrimination of positional isomers with >70% accuracy. Applied to binary glycan mixtures spanning structurally distinct pairs to positional isomers, DynaSpec-guided spectral windows enable non-negative least-squares unmixing with R² > 0.99 for fucosylation and galactosylation differences, while benchmarking near-superimposable positional isomers. DynaSpec further generalizes computationally to O-glycan cores and glycosaminoglycans, with experimental validation against hyaluronic acid. Together, these capabilities establish an ensemble-resolved framework for structure-informed spectroscopy across diverse biomolecular classes.