TY - JOUR
T1 - Immobilization of Phosphatidylinositides Revealed by Bilayer Leaflet Decoupling
AU - Sun, Simou
AU - Liu, Chang
AU - Rodriguez Melendez, Danixa
AU - Yang, Tinglu
AU - Cremer, Paul S.
N1 - Funding Information:
The authors thank Dr. Djoshkun Shengjuler for help with the PLCδ1-PH domain expression and purification. This work was supported by the National Science Foundation (CHE-1709735).
Publisher Copyright:
Copyright © 2020 American Chemical Society.
PY - 2020/7/29
Y1 - 2020/7/29
N2 - Phosphatidylinositol 4,5-bisphosphate (PIP2) has a significantly lower mobile fraction than most other lipids in supported lipid bilayers (SLBs). Moreover, the fraction of mobile PIP2 continuously decreases with time. To explore this, a bilayer unzipping technique was designed to uncouple the two leaflets of the SLB. The results demonstrate that PIP2 molecules in the top leaflet are fully mobile, while the PIP2 molecules in the lower leaflet are immobilized on the oxide support. Over time, mobile PIP2 species flip from the top leaflet to the bottom leaflet and become trapped. It was found that PIP2 flipped between the leaflets through a defect-mediated process. The flipping could be completely inhibited when holes in the bilayer were backfilled with bovine serum albumin (BSA). Moreover, by switching from H2O to D2O, it was shown that the primary interaction between PIP2 and the underlying substrate was due to hydrogen bond formation, which outcompeted electrostatic repulsion. Using substrates with fewer surface silanol groups, like oxidized polydimethylsiloxane, led to a large increase in the mobile fraction of PIP2. Moreover, PIP2 immobilization also occurred when the bilayer was supported on a protein surface rather than glass. These results may help to explain the behavior of PIP2 on the inner leaflet of the plasma membrane, where it is involved in attaching the membrane to the underlying cytoskeleton.
AB - Phosphatidylinositol 4,5-bisphosphate (PIP2) has a significantly lower mobile fraction than most other lipids in supported lipid bilayers (SLBs). Moreover, the fraction of mobile PIP2 continuously decreases with time. To explore this, a bilayer unzipping technique was designed to uncouple the two leaflets of the SLB. The results demonstrate that PIP2 molecules in the top leaflet are fully mobile, while the PIP2 molecules in the lower leaflet are immobilized on the oxide support. Over time, mobile PIP2 species flip from the top leaflet to the bottom leaflet and become trapped. It was found that PIP2 flipped between the leaflets through a defect-mediated process. The flipping could be completely inhibited when holes in the bilayer were backfilled with bovine serum albumin (BSA). Moreover, by switching from H2O to D2O, it was shown that the primary interaction between PIP2 and the underlying substrate was due to hydrogen bond formation, which outcompeted electrostatic repulsion. Using substrates with fewer surface silanol groups, like oxidized polydimethylsiloxane, led to a large increase in the mobile fraction of PIP2. Moreover, PIP2 immobilization also occurred when the bilayer was supported on a protein surface rather than glass. These results may help to explain the behavior of PIP2 on the inner leaflet of the plasma membrane, where it is involved in attaching the membrane to the underlying cytoskeleton.
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U2 - 10.1021/jacs.0c03800
DO - 10.1021/jacs.0c03800
M3 - Article
C2 - 32687699
AN - SCOPUS:85089179856
SN - 0002-7863
VL - 142
SP - 13003
EP - 13010
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 30
ER -