Steric Pressure between Proteins Opposes Membrane Phase Separation

Scheve, Christine S.; Gonzales, Paul A.; Momin, Noor; Stachowiak, Jeanne C.

doi:10.1016/j.bpj.2012.11.276

Cited by 2 publications

(2 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…A1) that has been described previously. 42 This microscope was equipped with a 488 nm picosecond pulsed laser and a time-correlated single photon counting electronic card (Becker and Hickl). Imaging wells were coated with lipid bilayers consisting of 100% DOPC to prevent adsorption of protein from solution.…”

Section: Fluorescence Correlation Spectroscopymentioning

confidence: 99%

Molecular Mechanisms of Membrane Curvature Sensing by a Disordered Protein

et al. 2019

Self Cite

View full text Add to dashboard Cite

The ability of proteins to sense membrane curvature is essential for the initiation and assembly of curved membrane structures. Established mechanisms of curvature sensing rely on proteins with specific structural features. In contrast, it has recently been discovered that intrinsically disordered proteins, which lack a defined three-dimensional fold, can also be potent sensors of membrane curvature. How can an unstructured protein sense the structure of the membrane surface? Many disordered proteins that associate with membranes have two key physical features -a high degree of conformational entropy and a high net negative charge. Binding of such proteins to membrane surfaces results simultaneously in a decrease in conformational entropy and an increase in electrostatic repulsion by anionic lipids. Here we show that each of these effects gives rise to a distinct mechanism of curvature sensing. Specifically, as the curvature of the membrane increases, the steric hindrance between the disordered protein and membrane is reduced, leading to an increase in chain entropy. At the same time, increasing membrane curvature increases the average separation between anionic amino acids and lipids, creating an electrostatic preference for curved membranes. Using quantitative imaging of membrane vesicles, our results demonstrate that long disordered amino acid chains with low net charge sense curvature predominately through the entropic mechanism. In contrast, shorter, more highly charged amino acid chains rely largely on the electrostatic mechanism. These findings provide a roadmap for predicting and testing the curvature sensitivity of the large and diverse set of disordered proteins that function at cellular membranes.

show abstract

Section: Fluorescence Correlation Spectroscopymentioning

confidence: 99%

Molecular Mechanisms of Membrane Curvature Sensing by a Disordered Protein

et al. 2019

Self Cite

View full text Add to dashboard Cite

show abstract

“…The pRSET vector containing the nondimerizable hexa-His-tagged eGFP (hisGFP A206K) was provided by A. Arkin (University of California, Berkeley, CA). Following published purification protocols, 24 his-eGFP was expressed in BL21(DE3) pLysS cells overnight at 18 °C and purified from bacterial extracts by incubation with Ni-NTA agarose beads in 25 mM HEPES, 150 mM NaCl, 1 mM imidazole, and 1 mM β-mercaptoethanol (pH 7.4). After being extensively washed with 25 mM HEPES, 150−300 mM NaCl, 1−5 mM imidazole, and 1 mM β-mercaptoethanol (pH 7.4), proteins were eluted via a gradient imidazole wash to a final concentration of 250 mM.…”

mentioning

confidence: 99%

The Display of Single-Domain Antibodies on the Surfaces of Connectosomes Enables Gap Junction-Mediated Drug Delivery to Specific Cell Populations

et al. 2017

Self Cite

View full text Add to dashboard Cite

Gap junctions, transmembrane protein channels that directly connect the cytoplasm of neighboring cells and enable the exchange of molecules between cells, are a promising new frontier for therapeutic delivery. Specifically, cell-derived lipid vesicles that contain functional gap junction channels, termed Connectosomes, have recently been demonstrated to substantially increase the effectiveness of small molecule chemotherapeutics. However, because gap junctions are present in nearly all tissues, Connectosomes have no intrinsic ability to target specific cell types, which potentially limits their therapeutic effectiveness. To address this challenge, here we display targeting ligands consisting of single-domain antibodies on the surfaces of Connectosomes. We demonstrate that these targeted Connectosomes selectively interact with cells that express a model receptor, promoting the selective delivery of the chemotherapeutic doxorubicin to this target cell population. More generally, our approach has the potential to boost cytoplasmic delivery of diverse therapeutic molecules to specific cell populations while protecting off-target cells, a critical step toward realizing the therapeutic potential of gap junctions.

show abstract

Steric Pressure between Proteins Opposes Membrane Phase Separation

Cited by 2 publications

References 0 publications

Molecular Mechanisms of Membrane Curvature Sensing by a Disordered Protein

Molecular Mechanisms of Membrane Curvature Sensing by a Disordered Protein

The Display of Single-Domain Antibodies on the Surfaces of Connectosomes Enables Gap Junction-Mediated Drug Delivery to Specific Cell Populations

Contact Info

Product

Resources

About