Spectroscopic and molecular-level characterization of partricins' self-association in aqueous solution

Aromatic heptaene macrolides are structurally related to amphotericin B, an agent of reference in treatment of systemic fungal infections, but contain an additional aromatic side chain and a native cis-trans polyene chromophore. Their strong biological activity and toxicity are likely influenced not only by molecular structure, but also by supramolecular organization in aqueous media. Here, we combined concentration-dependent UV-Vis and circular dichroism (CD) spectroscopy with molecular simulations to characterize the self-association of native partricins and their all-trans photoisomers. PCA-assisted spectral decomposition showed that all four antibiotics undergo self-association in phosphate buffer over the investigated concentration range. Both UV-Vis and CD data were best described by an effective two-state nucleation-elongation model, with subsequent aggregate growth favored over initial association. UV-Vis indicated broadly comparable transition concentrations across the series, whereas CD was more sensitive to chiral supramolecular organization. For the all-trans isoforms, absorption-detected association preceded the emergence of the dominant CD-active state. Umbrella-sampling simulations showed favorable pairwise association at short interchromophore distances for all compounds, with all-trans isomerization preserving the preferred association distance while slightly stabilizing the associated state. Two-dimensional well-tempered metadynamics further demonstrated that these distance-defined dimers are orientationally heterogeneous and include near-periplanar, antiparallel-like, and intermediate chromophore arrangements, with isoParB showing the most distinct orientational landscape. Unbiased multi-molecule simulations confirmed spontaneous aggregation beyond dimer formation and revealed dynamic growth of higher-order assemblies, particularly for isoParB. Overall, the results support a hierarchical model in which partricins form favorable, orientationally diverse low-order contacts that evolve into larger supramolecular aggregates. All-trans isomerization modulates this aggregation landscape without abolishing the intrinsic tendency toward self-association.