Nanoscale Patterning of Membrane-Bound Proteins Formed through Curvature-Induced Partitioning of Phase-Specific Receptor Lipids

Ogunyankin, Maria O.; Huber, Dale L.; Sasaki, Darryl Y.; Longo, Marjorie L.

doi:10.1021/la401011d

Cited by 14 publications

(14 citation statements)

References 37 publications

(52 reference statements)

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“…The demonstration of PLM performed here measured nanoscale hemispherical membrane curvature of an SLB draped over NPs ranging in radii from 24 to 70 nm. Prior methods of inducing nanoscale curvature utilized nanoengineered wavy glass substrates (47), microfabricated structures (48,49), membrane tubule pulled from GUVs (50), and SLBs on deformable substrates (51,52). However, wavy glass substrates, thick polymer structures, and lipid tubules are not compatible with TIRF excitation.…”

Section: Membrane Topography Over Npsmentioning

confidence: 99%

The Detection of Nanoscale Membrane Bending with Polarized Localization Microscopy

Kabbani

Kelly

2017

Biophysical Journal

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The curvature of biological membranes at the nanometer scale is critically important for vesicle trafficking, organelle morphology, and disease propagation. The initial membrane bending events that initiate these complex processes occur at a length scale that is irresolvable by most super-resolution optical microscopy methods. This manuscript reports the development of polarized localization microscopy (PLM), a pointillist optical imaging technique for the detection of nanoscale membrane curvature in correlation with single-molecule dynamics and molecular sorting. PLM combines polarized total internal reflection fluorescence microscopy (TIRFM) and single-molecule localization microscopy to reveal membrane orientation with a sub-diffractionlimited resolution without reducing localization precision by point spread function (PSF) manipulation. Membrane curvature detection with PLM requires fewer localization events to detect curvature than 3D single-molecule localization microscopy (e.g., PALM or STORM), which enables curvature detection 10x faster via PLM. With rotationally confined lipophilic fluorophores and the polarized incident fluorescence excitation, membrane-bending events are revealed with super-resolution. Engineered hemispherical membrane curvature with a radii ≥ 24 nm was detected with PLM and individual fluorophore localization precision was 13 ± 5 nm.Further, deciphering molecular mobility as a function of membrane topology was enabled. The diffusion coefficient of individual DiI molecules was 7.6x higher in planar supported lipid bilayers than within nanoscale membrane curvature. Through the theoretical foundation and experimental demonstration provided here, PLM is poised to become a powerful technique for revealing the underlying biophysical mechanisms of membrane bending at physiological length scales.

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Section: Membrane Topography Over Npsmentioning

confidence: 99%

The Detection of Nanoscale Membrane Bending with Polarized Localization Microscopy

Kabbani

Kelly

2017

Biophysical Journal

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“…Although PDMS is used widely in soft-lithography, the temporal stability of phospholipid bilayers on plasma-oxidized PDMS (ox-PDMS) is not known. Such a study is useful, particularly since the use of ox-PDMS as a biointerface, instead of glass, is advantageous in applications such as microfluidics[6], applications that require variations in topography and curvature[8,10,11,14,15,30], and applications that require dynamic changes in surface area[14,19]. …”

Section: Introductionmentioning

confidence: 99%

Lipid Bilayers Are Long-Lived on Solvent Cleaned Plasma-Oxidized poly(dimethyl)siloxane (ox-PDMS)

et al. 2017

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Although it is well known that phospholipids self-assemble on hydrophilic plasma-oxidized PMDS surfaces (ox-PDMS) to form cell membrane mimetic bilayers, the temporal stability of phospholipid membranes on these surfaces is unknown. Here we report that phospholipid bilayers remain stable on solvent-cleaned ox-PDMS for at least 132 hours after preparation. Absent solvent cleaning, the bilayers were stable for only 36 hours. We characterized the phospholipid bilayers, i) through quantitative comparative analysis of the fluorescence intensity of phospholipid bilayers on ox-PDMS and phospholipid monolayers on native PDMS and, ii) through measurements of the diffusive mobility of the lipids through fluorescence recovery after photobleaching (FRAP). The fluorescence intensity of the phospholipid layer remained consistent with that of a bilayer for 132 hours. The evolution of the diffusive mobility of the phospholipids in the bilayer on ox-PDMS over time was similar to lipids in control bilayers prepared on glass surfaces. Solvent cleaning was essential for the long-term stability of the bilayers on ox-PDMS. Without cleaning in acetone and isopropanol, phospholipid bilayers prepared on ox-PDMS surfaces peeled off in large patches within 36 hours. Importantly, we find that phospholipid bilayers supported on solvent-cleaned ox-PDMS were indistinguishable from phospholipid bilayers supported on glass for at least 36 hours after preparation. Our results provide a link between the two common surfaces used to prepare in vitro biomimetic phospholipid membranes—i) glass surfaces used predominantly in fundamental biophysical experiments, for which there is abundant physicochemical information, with ii) ox-PDMS, the dominant material used in practical, applications-oriented systems to build micro-devices, topographically-patterned surfaces, and biosensors where there is a dearth of information.

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“…Phase-specific adsorption and binding of proteins has been of interest for the development of biomaterials for various applications, such as small-scale fluidic devices, biosensors, and high-density arrays. 8–12 …”

Section: Introductionmentioning

confidence: 99%

Crowding-Induced Mixing Behavior of Lipid Bilayers: Examination of Mixing Energy, Phase, Packing Geometry, and Reversibility

Zeno¹,

Rystov²,

Sasaki

et al. 2016

Langmuir

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In an effort to develop a general thermodynamic model from first principles to describe mixing behavior of lipid membranes we examined lipid mixing induced by targeted binding of small (Green Fluorescent Protein (GFP)) and large (Nanolipoprotein Particles (NLPs)) structures to specific phases of phase separated lipid bilayers. Phases were targeted by incorporation of phase-partitioning iminodiacetic acid (IDA) functionalized lipids into ternary lipid mixtures consisting of DPPC, DOPC and cholesterol. GFP and NLPs, containing histidine tags, bound the IDA portion of these lipids via a metal, Cu2+, chelating mechanism. In giant unilamellar vesicles (GUVs), GFP and NLPs bound to the Lo domains of bilayers containing DPIDA, and bound to the Ld region of bilayers containing DOIDA. At sufficiently large concentrations of DPIDA or DOIDA, lipid mixing was induced by bound GFP and NLPs. The validity of the thermodynamic model was confirmed when it was found that the statistical mixing distribution as a function of crowding energy for smaller GFP and larger NLPs collapsed to the same trend line for each GUV composition. Moreover, results of this analysis show that the free energy of mixing for a ternary lipid bilayer consisting of DOPC, DPPC, and cholesterol varied from 7.9×10−22 to 1.5×10−20 Joules/lipid at the compositions observed, decreasing as the relative cholesterol concentration was increased. It was discovered that there appears to be a maximum packing density, and associated maximum crowding pressure, of the NLPs, suggestive of circular packing. A similarity in mixing induced by NLP1 and NLP3 despite large difference in projected areas was analytically consistent with monovalent (one histidine) vs. divalent (two histidine) surface interactions, respectively. In addition to GUVs, binding and induced mixing behavior of NLPs was also observed on planar, supported lipid multibilayers. The mixing process was reversible, with Lo domains reappearing after addition of EDTA for NLP removal.

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Nanoscale Patterning of Membrane-Bound Proteins Formed through Curvature-Induced Partitioning of Phase-Specific Receptor Lipids

Cited by 14 publications

References 37 publications

The Detection of Nanoscale Membrane Bending with Polarized Localization Microscopy

The Detection of Nanoscale Membrane Bending with Polarized Localization Microscopy

Lipid Bilayers Are Long-Lived on Solvent Cleaned Plasma-Oxidized poly(dimethyl)siloxane (ox-PDMS)

Crowding-Induced Mixing Behavior of Lipid Bilayers: Examination of Mixing Energy, Phase, Packing Geometry, and Reversibility

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