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Activation of bacterial channel MscL in mechanically stimulated droplet interface bilayers.

Najem JS, Dunlap MD, Rowe ID, Freeman EC, Grant JW, Sukharev S, Leo DJ - Sci Rep (2015)

Bottom Line: Geometrical analysis of droplets during compression indicates that both contact angle and total area of the water-oil interfaces contribute to the generation of tension in the bilayer.The measured expansion of the interfaces by 2.5% is predicted to generate a 4-6 mN/m tension in the bilayer, just sufficient for gating.This work clarifies the principles of interconversion between bulk and surface forces in the DIB, facilitates the measurements of fundamental membrane properties, and improves our understanding of MscL response to membrane tension.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, United States.

ABSTRACT
MscL, a stretch-activated channel, saves bacteria experiencing hypo-osmotic shocks from lysis. Its high conductance and controllable activation makes it a strong candidate to serve as a transducer in stimuli-responsive biomolecular materials. Droplet interface bilayers (DIBs), flexible insulating scaffolds for such materials, can be used as a new platform for incorporation and activation of MscL. Here, we report the first reconstitution and activation of the low-threshold V23T mutant of MscL in a DIB as a response to axial compressions of the droplets. Gating occurs near maximum compression of both droplets where tension in the membrane is maximal. The observed 0.1-3 nS conductance levels correspond to the V23T-MscL sub-conductive and fully open states recorded in native bacterial membranes or liposomes. Geometrical analysis of droplets during compression indicates that both contact angle and total area of the water-oil interfaces contribute to the generation of tension in the bilayer. The measured expansion of the interfaces by 2.5% is predicted to generate a 4-6 mN/m tension in the bilayer, just sufficient for gating. This work clarifies the principles of interconversion between bulk and surface forces in the DIB, facilitates the measurements of fundamental membrane properties, and improves our understanding of MscL response to membrane tension.

No MeSH data available.


Related in: MedlinePlus

Changes in droplet areas and contact angles revealed by imaging and area calculations in the course of cyclic mechanical stimulation.(a) The zoomed-in view of the one droplet and determination of the entire interfacial area, the bilayer area, and the contact angle. (b) The bilayer angle as a function of the displacement. (c) The relative area changes as a function of frequency of oscillation.
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f7: Changes in droplet areas and contact angles revealed by imaging and area calculations in the course of cyclic mechanical stimulation.(a) The zoomed-in view of the one droplet and determination of the entire interfacial area, the bilayer area, and the contact angle. (b) The bilayer angle as a function of the displacement. (c) The relative area changes as a function of frequency of oscillation.

Mentions: The equation above suggests that γb is always lower than 2γm, and for this to be true, the Plateau-Gibbs border must move away from the centre of the bilayer in order to increase γb. Note that at the Plateau-Gibbs border, the monolayers (water-oil interface) are near spherical according to the Young-Laplace equation22. Therefore, under conditions of constant volume, the area of the monolayer covering the entire water-oil interface in both compressed droplets increases. The relative expansion of the droplets is measured at different frequencies while maintaining a 100 mV transmembrane potential. Our results (Fig. 7c) show that at 0.2 Hz the droplet relative area change is approximately 2.5%. However, as we increased the frequency of oscillation to 7.5 Hz this value dropped to approximately 0.1%. The elastic modulus for isothermal area compression or extension is defined in terms of the mechanical properties of the monolayer as follows:


Activation of bacterial channel MscL in mechanically stimulated droplet interface bilayers.

Najem JS, Dunlap MD, Rowe ID, Freeman EC, Grant JW, Sukharev S, Leo DJ - Sci Rep (2015)

Changes in droplet areas and contact angles revealed by imaging and area calculations in the course of cyclic mechanical stimulation.(a) The zoomed-in view of the one droplet and determination of the entire interfacial area, the bilayer area, and the contact angle. (b) The bilayer angle as a function of the displacement. (c) The relative area changes as a function of frequency of oscillation.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4562232&req=5

f7: Changes in droplet areas and contact angles revealed by imaging and area calculations in the course of cyclic mechanical stimulation.(a) The zoomed-in view of the one droplet and determination of the entire interfacial area, the bilayer area, and the contact angle. (b) The bilayer angle as a function of the displacement. (c) The relative area changes as a function of frequency of oscillation.
Mentions: The equation above suggests that γb is always lower than 2γm, and for this to be true, the Plateau-Gibbs border must move away from the centre of the bilayer in order to increase γb. Note that at the Plateau-Gibbs border, the monolayers (water-oil interface) are near spherical according to the Young-Laplace equation22. Therefore, under conditions of constant volume, the area of the monolayer covering the entire water-oil interface in both compressed droplets increases. The relative expansion of the droplets is measured at different frequencies while maintaining a 100 mV transmembrane potential. Our results (Fig. 7c) show that at 0.2 Hz the droplet relative area change is approximately 2.5%. However, as we increased the frequency of oscillation to 7.5 Hz this value dropped to approximately 0.1%. The elastic modulus for isothermal area compression or extension is defined in terms of the mechanical properties of the monolayer as follows:

Bottom Line: Geometrical analysis of droplets during compression indicates that both contact angle and total area of the water-oil interfaces contribute to the generation of tension in the bilayer.The measured expansion of the interfaces by 2.5% is predicted to generate a 4-6 mN/m tension in the bilayer, just sufficient for gating.This work clarifies the principles of interconversion between bulk and surface forces in the DIB, facilitates the measurements of fundamental membrane properties, and improves our understanding of MscL response to membrane tension.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, United States.

ABSTRACT
MscL, a stretch-activated channel, saves bacteria experiencing hypo-osmotic shocks from lysis. Its high conductance and controllable activation makes it a strong candidate to serve as a transducer in stimuli-responsive biomolecular materials. Droplet interface bilayers (DIBs), flexible insulating scaffolds for such materials, can be used as a new platform for incorporation and activation of MscL. Here, we report the first reconstitution and activation of the low-threshold V23T mutant of MscL in a DIB as a response to axial compressions of the droplets. Gating occurs near maximum compression of both droplets where tension in the membrane is maximal. The observed 0.1-3 nS conductance levels correspond to the V23T-MscL sub-conductive and fully open states recorded in native bacterial membranes or liposomes. Geometrical analysis of droplets during compression indicates that both contact angle and total area of the water-oil interfaces contribute to the generation of tension in the bilayer. The measured expansion of the interfaces by 2.5% is predicted to generate a 4-6 mN/m tension in the bilayer, just sufficient for gating. This work clarifies the principles of interconversion between bulk and surface forces in the DIB, facilitates the measurements of fundamental membrane properties, and improves our understanding of MscL response to membrane tension.

No MeSH data available.


Related in: MedlinePlus