Details

Autor(en) / Beteiligte

• Tei, Reika
• Bagde, Saket R
• Fromme, J Christopher
• Baskin, Jeremy M

Titel

Activity-based directed evolution of a membrane editor in mammalian cells

Ist Teil von

• Nature chemistry, 2023-07, Vol.15 (7), p.1030

Ort / Verlag

England

Erscheinungsjahr

2023

Quelle

MEDLINE

Beschreibungen/Notizen

• Cellular membranes contain numerous lipid species, and efforts to understand the biological functions of individual lipids have been stymied by a lack of approaches for controlled modulation of membrane composition in situ. Here we present a strategy for editing phospholipids, the most abundant lipids in biological membranes. Our membrane editor is based on a bacterial phospholipase D (PLD), which exchanges phospholipid head groups through hydrolysis or transphosphatidylation of phosphatidylcholine with water or exogenous alcohols. Exploiting activity-dependent directed enzyme evolution in mammalian cells, we have developed and structurally characterized a family of 'superPLDs' with up to a 100-fold enhancement in intracellular activity. We demonstrate the utility of superPLDs for both optogenetics-enabled editing of phospholipids within specific organelle membranes in live cells and biocatalytic synthesis of natural and unnatural designer phospholipids in vitro. Beyond the superPLDs, activity-based directed enzyme evolution in mammalian cells is a generalizable approach to engineer additional chemoenzymatic biomolecule editors.