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Regulation of PLCβ2 by the electrostatic and mechanical properties of lipid bilayers.

Arduin A, Gaffney PR, Ces O - Sci Rep (2015)

Bottom Line: However, the regulatory mechanism of PLC is not yet understood in detail.Evidence was found for a direct interaction between PLC and the GTPases that mediate phospholipase activation.PLC activity was found to depend upon the electrostatic potential and the stored curvature elastic stress of the lipid membranes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Chemical Biology, Department of Chemical Biology, Imperial College London, Exhibition Road, London SW7 2AZ, United Kingdom.

ABSTRACT
Phosphoinositide-specific phospholipase C (PLC) is an important family of enzymes constituting a junction between phosphoinositide lipid signaling and the trans-membrane signal transduction processes that are crucial to many living cells. However, the regulatory mechanism of PLC is not yet understood in detail. To address this issue, activity studies were carried out using lipid vesicles in a model system that was specifically designed to study protein-protein and lipid-protein interactions in concert. Evidence was found for a direct interaction between PLC and the GTPases that mediate phospholipase activation. Furthermore, for the first time, the relationships between PLC activity and substrate presentation in lipid vesicles of various sizes, as well as lipid composition and membrane mechanical properties, were analyzed. PLC activity was found to depend upon the electrostatic potential and the stored curvature elastic stress of the lipid membranes.

No MeSH data available.


Related in: MedlinePlus

PLCβ2 activity measurements obtained using the cell-free system.The bar chart shows the average activity after sample incubation at 30 °C for 45 min, measured in duplicate samples (± the s.e.m.). The samples contained: purified w.t. PLCβ2 (2–803 AA) (0.25 ng/μl), or purified PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl), and DOPE: PtdIns (4,5)P2/[3H]-PtdIns(4,5)P2 lipid vesicles 16:1 mol/mol. All other conditions are the same as for Fig. 2.
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f9: PLCβ2 activity measurements obtained using the cell-free system.The bar chart shows the average activity after sample incubation at 30 °C for 45 min, measured in duplicate samples (± the s.e.m.). The samples contained: purified w.t. PLCβ2 (2–803 AA) (0.25 ng/μl), or purified PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl), and DOPE: PtdIns (4,5)P2/[3H]-PtdIns(4,5)P2 lipid vesicles 16:1 mol/mol. All other conditions are the same as for Fig. 2.

Mentions: We tested different concentrations of PLCβ2 (2–803 AA) ∆(470–540), in which the X-Y linker had been deleted, to find values that gave a similar response to w.t. PLCβ2 (2–803 AA) in the cell-free system. In the presence of GTPγS and GG-Rac2, only 1.5 nM concentration of PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl) was required to obtain a stimulation response similar to that from 2.7 nM w.t. PLCβ2 (2–803 AA) (0.25 ng/μl) (Fig. 9, columns 2 and 4). Furthermore, in the presence of GDP and GG-Rac2, the activity of w.t. PLCβ2 (2–803 AA) was lower than the activity of PLCβ2 (2–803 AA) ∆(470–540) (Fig. 9, columns 1 and 3).


Regulation of PLCβ2 by the electrostatic and mechanical properties of lipid bilayers.

Arduin A, Gaffney PR, Ces O - Sci Rep (2015)

PLCβ2 activity measurements obtained using the cell-free system.The bar chart shows the average activity after sample incubation at 30 °C for 45 min, measured in duplicate samples (± the s.e.m.). The samples contained: purified w.t. PLCβ2 (2–803 AA) (0.25 ng/μl), or purified PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl), and DOPE: PtdIns (4,5)P2/[3H]-PtdIns(4,5)P2 lipid vesicles 16:1 mol/mol. All other conditions are the same as for Fig. 2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4525138&req=5

f9: PLCβ2 activity measurements obtained using the cell-free system.The bar chart shows the average activity after sample incubation at 30 °C for 45 min, measured in duplicate samples (± the s.e.m.). The samples contained: purified w.t. PLCβ2 (2–803 AA) (0.25 ng/μl), or purified PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl), and DOPE: PtdIns (4,5)P2/[3H]-PtdIns(4,5)P2 lipid vesicles 16:1 mol/mol. All other conditions are the same as for Fig. 2.
Mentions: We tested different concentrations of PLCβ2 (2–803 AA) ∆(470–540), in which the X-Y linker had been deleted, to find values that gave a similar response to w.t. PLCβ2 (2–803 AA) in the cell-free system. In the presence of GTPγS and GG-Rac2, only 1.5 nM concentration of PLCβ2 (2–803 AA) ∆(470–540) (0.125 ng/μl) was required to obtain a stimulation response similar to that from 2.7 nM w.t. PLCβ2 (2–803 AA) (0.25 ng/μl) (Fig. 9, columns 2 and 4). Furthermore, in the presence of GDP and GG-Rac2, the activity of w.t. PLCβ2 (2–803 AA) was lower than the activity of PLCβ2 (2–803 AA) ∆(470–540) (Fig. 9, columns 1 and 3).

Bottom Line: However, the regulatory mechanism of PLC is not yet understood in detail.Evidence was found for a direct interaction between PLC and the GTPases that mediate phospholipase activation.PLC activity was found to depend upon the electrostatic potential and the stored curvature elastic stress of the lipid membranes.

View Article: PubMed Central - PubMed

Affiliation: Institute of Chemical Biology, Department of Chemical Biology, Imperial College London, Exhibition Road, London SW7 2AZ, United Kingdom.

ABSTRACT
Phosphoinositide-specific phospholipase C (PLC) is an important family of enzymes constituting a junction between phosphoinositide lipid signaling and the trans-membrane signal transduction processes that are crucial to many living cells. However, the regulatory mechanism of PLC is not yet understood in detail. To address this issue, activity studies were carried out using lipid vesicles in a model system that was specifically designed to study protein-protein and lipid-protein interactions in concert. Evidence was found for a direct interaction between PLC and the GTPases that mediate phospholipase activation. Furthermore, for the first time, the relationships between PLC activity and substrate presentation in lipid vesicles of various sizes, as well as lipid composition and membrane mechanical properties, were analyzed. PLC activity was found to depend upon the electrostatic potential and the stored curvature elastic stress of the lipid membranes.

No MeSH data available.


Related in: MedlinePlus