2016
DOI: 10.7554/elife.17438
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The dimerization equilibrium of a ClC Cl−/H+ antiporter in lipid bilayers

Abstract: Interactions between membrane protein interfaces in lipid bilayers play an important role in membrane protein folding but quantification of the strength of these interactions has been challenging. Studying dimerization of ClC-type transporters offers a new approach to the problem, as individual subunits adopt a stable and functionally verifiable fold that constrains the system to two states – monomer or dimer. Here, we use single-molecule photobleaching analysis to measure the probability of ClC-ec1 subunit ca… Show more

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Cited by 53 publications
(137 citation statements)
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References 51 publications
(81 reference statements)
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“…CLC-ec1 C85A/H234C including the hexahistidine tag has a molecular weight of 51,997 g/mole, MW POPE = 717.996 g/mole and MW POPG = 770.989 g/mole. Therefore, a proteoliposome sample reconstituted at 1 µg/mg in 2:1 POPE/POPG lipids corresponds to a mole fraction of: Assuming a 50% mole fraction recovery as was previously observed (14), then our observed mole fraction is χ observed = 0.7 x 10 −5 subunits/lipid.…”
Section: Methodssupporting
confidence: 70%
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“…CLC-ec1 C85A/H234C including the hexahistidine tag has a molecular weight of 51,997 g/mole, MW POPE = 717.996 g/mole and MW POPG = 770.989 g/mole. Therefore, a proteoliposome sample reconstituted at 1 µg/mg in 2:1 POPE/POPG lipids corresponds to a mole fraction of: Assuming a 50% mole fraction recovery as was previously observed (14), then our observed mole fraction is χ observed = 0.7 x 10 −5 subunits/lipid.…”
Section: Methodssupporting
confidence: 70%
“…MATLAB was used to simulate the random process of multi-species subunit capture into a heterogeneous liposome population, as described previously (14, 18). For each liposome sub-population with radius r , a matrix was created with size N liposomes (r) , and each N protein (r) sub-species was randomly assigned to the liposome population.…”
Section: Methodsmentioning
confidence: 99%
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“…During incorporation, the Cy-ncAA may be exposed to reductive conditions in the cell that lead to formation of dark state adducts (Dempsey et al, 2009). Furthermore, imaging was performed in the absence of typical oxygen scavenger systems, as this has been shown to lead to improved photo-bleaching traces (Chadda et al, 2016) but could reduce the overall fluorescent yield. In this regard, future encoding experiments performed in the presence of oxygen quenchers or more stable fluorophores (Zheng et al, 2014) may be required for specific applications of the approach.…”
Section: Resultsmentioning
confidence: 99%