Phosphatidylserine externalization on the surface of dying cells is a key signal for their recognition and clearance by macrophages and is mediated by members of the X-Kell related (Xkr) protein family. Defective Xkr-mediated scrambling impairs clearance, leading to inflammation. It was proposed that activation of the Xkr4 apoptotic scramblase requires caspase cleavage, followed by dimerization and ligand binding. Here, using a combination of biochemical approaches we show that purified monomeric, full-length human Xkr4 (hXkr4) scrambles lipids. CryoEM imaging shows that hXkr4 adopts a novel conformation, where three conserved acidic residues create a negative electrostatic surface embedded in the membrane. Molecular dynamics simulations show this conformation induces membrane thinning, which could promote scrambling. Thinning is ablated or reduced in conditions where scrambling is abolished or reduced. Our work provides insights into the molecular mechanisms of hXkr4 scrambling and su

Xkr4 apoptotic scramblase activation is thought to involve caspase cleavage and dimerization to expose phosphatidylserine on dying cells. The authors show that full-length Xkr4 is an active monomeric ...

To test whether Xkr proteins are scramblases we sought to purify and functionally reconstitute them in proteoliposomes. A screen of GFP-tagged family members using fluorescence size exclusion chromatography (FSEC)44,45 identified human Xkr4 (hXkr4, MW ~ 71 kDa) as a promising...