ja508631n_si_001.pdf (19.16 MB)
Quantifying Transient Interactions between Bacillus Phosphatidylinositol-Specific Phospholipase‑C and Phosphatidylcholine-Rich Vesicles
journal contribution
posted on 2015-12-22, 22:32 authored by Boqian Yang, Mingming Pu, Hanif M. Khan, Larry Friedman, Nathalie Reuter, Mary F. Roberts, Anne GershensonBacillus 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.