title: Quantifying transient interactions between Bacillus phosphatidylinositol-specific phospholipase-C and phosphatidylcholine-rich vesicles.
abstract: Bacillus thuringiensis secretes the virulence factor phosphatidylinositol-specific phospholipase C (BtPI-PLC), which specifically binds to phosphatidylcholine (PC) and cleaves GPI-anchored proteins off eukaryotic plasma membranes. To elucidate how BtPI-PLC searches for GPI-anchored proteins on the membrane surface, we measured residence times of single fluorescently labeled proteins on PC-rich small unilamellar vesicles (SUVs). BtPI-PLC interactions with the SUV surface are transient with a lifetime of 379 ± 49 ms. These data also suggest that BtPI-PLC does not directly sense curvature, but rather prefers to bind to the numerous lipid packing defects in SUVs. Despite this preference for defects, all-atom molecular dynamics simulations of BtPI-PLC interacting with PC-rich bilayers show that the protein is shallowly anchored with the deepest insertions ∼18 Å above the bilayer center. Membrane partitioning is mediated, on average, by 41 hydrophobic, 8 hydrogen-bonding, and 2 cation-π (between PC choline headgroups and Tyr residues) transient interactions with phospholipids. These results lead to a quantitative model for BtPI-PLC interactions with cell membranes where protein binding is mediated by lipid packing defects, possibly near GPI-anchored proteins, and the protein diffuses on the membrane for ∼100-380 ms, during which time it may cleave ∼10 GPI-anchored proteins before dissociating. This combination of short two-dimensional scoots followed by three-dimensional hops may be an efficient search strategy on two-dimensional surfaces with obstacles.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Massachusetts , Amherst, Massachusetts 01003, United States.
authors.forename: Boqian
authors.initials: B
authors.lastname: Yang
authors.forename: Mingming
authors.initials: M
authors.lastname: Pu
authors.forename: Hanif M
authors.initials: HM
authors.lastname: Khan
authors.forename: Larry
authors.initials: L
authors.lastname: Friedman
authors.forename: Nathalie
authors.initials: N
authors.lastname: Reuter
authors.forename: Mary F
authors.initials: MF
authors.lastname: Roberts
authors.forename: Anne
authors.initials: A
authors.lastname: Gershenson
mesh_terms: D001413:Bacillus thuringiensis / Q000201:enzymology*; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D010713:Phosphatidylcholines / Q000378:metabolism*; D054801:Phosphoinositide Phospholipase C / Q000737:chemistry* / Q000378:metabolism*; D053835:Unilamellar Liposomes / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D010713:Phosphatidylcholines; D053835:Unilamellar Liposomes; D054801:Phosphoinositide Phospholipase C