Directed evolution of LaccID for cell surface proximity labeling and electron microscopy.

ABSTRACT: Enzymes that oxidize aromatic substrates have been harnessed for cell-based technologies including proximity labeling and electron microscopy; however, they are associated with drawbacks such as the need for toxic H₂O₂. Here, we explore multicopper oxidases (laccases) as a new enzyme class for proximity labeling and electron microscopy in mammalian cells. LaccID was generated through 11 rounds of directed evolution from an ancestral fungal laccase and catalyzes one-electron oxidation of diverse aromatic substrates using O₂ instead of toxic H₂O₂. Surprisingly, we found that LaccID is selectively active at the surface plasma membrane of both living and fixed cells. We use LaccID proximity labeling and mass spectrometry to map the changin

SUBMITTER: Lee SY 

PROVIDER: S-EPMC12643941 | biostudies-literature | 2025 Dec

REPOSITORIES: biostudies-literature