A Chemoproteomics Approach to Profile Phospholipase D‑Derived Phosphatidyl Alcohol Interactions

Alcohol
consumption leads to formation of phosphatidylethanol (PEth)
via the transphosphatidylation activity of phospholipase D (PLD) enzymes.
Though this non-natural phospholipid routinely serves as a biomarker
of chronic alcoholism, its pathophysiological roles remain unknown.
We use a minimalist diazirine alkyne alcohol as an ethanol surrogate
to generate clickable, photoaffinity lipid reporters of PEth localization
and lipid–protein interactions via PLD-mediated transphosphatidylation.
We use these tools to visualize phosphatidyl alcohols in a manner
compatible with standard permeabilization and immunofluorescence methods.
We also use click chemistry tagging, enrichment, and proteomics analysis
to define the phosphatidyl alcohol interactome. Our analysis reveals
an enrichment of putative interactors at various membrane locations,
and we validate one such interaction with the single-pass transmembrane
protein basigin/CD147. This study provides a comprehensive view of
the molecular interactions of phosphatidyl alcohols with the cellular
proteome and points to future work to connect such interactions to
potential pathophysiological roles of PEth.