Employing Metadynamics to Predict the Membrane Partitioning of Carboxy-2 <i>H</i> -Azirine Natural Products

Natural products containing the carboxy-2<i>H</i>-azirine moiety are an exciting target for investigation due to their broad-spectrum antimicrobial activity and new chemical space they afford for novel therapeutic development. The carboxy-2<i>H</i>-azirine moiety, including those appended to well-characterized chemical scaffolds, is understudied, which creates a challenge for understanding potential modes of inhibition. In particular, some known natural product carboxy-2<i>H</i>-azirines have long hydrophobic tails, which could implicate them in membrane-associated processes. In this study, we examined a small set of carboxy-2<i>H</i>-azirine natural products with varied structural features that could alter membrane partitioning. We compared the predicted membrane partitioning and alignment of these compounds to those of established membrane embedders with similar chemical scaffolds. To accomplish this, we developed parameters within the framework of the CHARMM36 force field for the 2<i>H</i>-azirine functional group and performed metadynamics simulations of the partitioning into a model bacterial membrane from aqueous solution. We determined that the carboxy-2<i>H</i>-azirine functional group is strongly hydrophilic, imbuing the long-chain natural products with amphipathicity similar to the known membrane-embedding molecules to which they were compared. For the long-chain analogs, the carboxy-2<i>H</i>-azirine head group stays within 1 nm of the phosphate layer, while the hydrophobic tail sits within the membrane. The carboxy-2<i>H</i>-azirine lacking the long alkyl chain instead partitions completely into aqueous solution.