A single gD glycoprotein can mediate infection by Herpes simplex virus.
Journal of the American Chemical Society 135(30):11175 (2013)
Herpes simplex viruses display hundreds of gD glycoproteins, and yet their neutralization requires tens of thousands of antibodies per virion, leading us to ask whether a wild-type virion with just a single free gD is still infective. By quantitative analysis of fluorescently labeled virus parti...
Pipette-surface interaction: current enhancement and intrinsic force.
Journal of the American Chemical Society 135(1):322 (2013)
There is an intrinsic repulsion between glass and cell surfaces that allows noninvasive scanning ion conductance microscopy (SICM) of cells and which must be overcome in order to form the gigaseals used for patch clamping investigations of ion channels. However, the interactions of surfaces in p...
Hydrodynamic trapping of molecules in lipid bilayers.
PNAS 109(26):10328 (2012)
In this work we show how hydrodynamic forces can be used to locally trap molecules in a supported lipid bilayer (SLB). The method uses the hydrodynamic drag forces arising from a flow through a conical pipette with a tip radius of 1-1.5 μm, placed approximately 1 μm above the investigated SLB. T...
Direct observation of the interconversion of normal and toxic forms of α-synuclein.
Cell 149(5):1048 (2012)
Here, we use single-molecule techniques to study the aggregation of α-synuclein, the protein whose misfolding and deposition is associated with Parkinson's disease. We identify a conformational change from the initially formed oligomers to stable, more compact proteinase-K-resistant oligomers as...
Single molecule fluorescence under conditions of fast flow.
Analytical chemistry 84(1):179 (2012)
We have experimentally determined the optimal flow velocities to characterize or count single molecules by using a simple microfluidic device to perform two-color coincidence detection (TCCD) and single pair Förster resonance energy transfer (spFRET) using confocal fluorescence spectroscopy on m...
The extracellular chaperone clusterin sequesters oligomeric forms of the amyloid-β(1-40) peptide.
Nature Structural & Molecular Biology 19(1):79 (2012)
In recent genome-wide association studies, the extracellular chaperone protein, clusterin, has been identified as a newly-discovered risk factor in Alzheimer's disease. We have examined the interactions between human clusterin and the Alzheimer's disease-associated amyloid-β(1-40) peptide (Aβ(1-...
Single-molecule fluorescence coincidence spectroscopy and its application to resonance energy transfer.
ChemPhysChem 12(3):491 (2011)
The use of Förster resonance energy transfer (FRET) as a tool to study biomolecules has been greatly enhanced by new advances in single-molecule fluorescence (SMF) techniques. This has allowed new insights into the structure and dynamics of complex biomolecular machinery. However, there are stil...
Fluorescence coincidence spectroscopy for single-molecule fluorescence resonance energy-transfer measurements.
Analytical chemistry 80(22):8389 (2008)
Single-molecule fluorescence resonance energy transfer (FRET) is commonly used to probe different conformations and conformational dynamics of single biomolecules. However, the analysis of raw burst traces is not always straightforward. The presence of a "zero peak" and the skewness of peaks at ...
Direct characterization of amyloidogenic oligomers by single-molecule fluorescence.
PNAS 105(38):14424 (2008)
A key issue in understanding the pathogenic conditions associated with the aberrant aggregation of misfolded proteins is the identification and characterization of species formed during the aggregation process. Probing the nature of such species has, however, proved to be extremely challenging t...
Single-molecule level analysis of the subunit composition of the T cell receptor on live T cells.
PNAS 104(45):17662 (2007)
The T cell receptor (TCR) expressed on most T cells is a protein complex consisting of TCRalphabeta heterodimers that bind antigen and cluster of differentiation (CD) 3epsilondelta, epsilongamma, and zetazeta dimers that initiate signaling. A long-standing controversy concerns whether there is o...
Two-color fluorescence analysis of individual virions determines the distribution of the copy number of proteins in herpes simplex virus particles.
Biophysical Journal 93(4):1329 (2007)
We present a single virion method to determine absolute distributions of copy number in the protein composition of viruses and apply it to herpes simplex virus type 1. Using two-color coincidence fluorescence spectroscopy, we determine the virion-to-virion variability in copy numbers of fluoresc...
Surface conductivity of biological macromolecules measured by nanopipette dielectrophoresis.
Physical Review Letters 98(19):198102 (2007)
We report the measurement of the surface conductivity of biological macromolecules by dielectrophoretic trapping at the tip of a glass nanopipet. We find that the threshold voltage for trapping is a function of salt concentration and can be directly linked to the effective conductivity of the bi...
Optimized threshold selection for single-molecule two-color fluorescence coincidence spectroscopy.
Analytical chemistry 79(7):2771 (2007)
Two-color coincidence detection is a single-molecule fluorescence technique that is capable of resolving subpopulations of biomolecular complexes at very low concentrations. In this paper, we have developed a method that automatically determines appropriate thresholds for the analysis of sets of...
A renewable nanosensor based on a glass nanopipette.
Journal of the American Chemical Society 128(51):16462 (2006)
A fluorescent nanosensor based on reporter dye molecules trapped in the tip of a nanopipette has been developed. This 100 nm sized nanosensor has been shown to be capable of measuring local pH and mapping sodium concentration with a temporal resolution of a few milliseconds.
Analysis of human telomerase activity and function by two color single molecule coincidence fluorescence spectroscopy.
Journal of the American Chemical Society 128(15):4992 (2006)
Telomerase is a nonclassical DNA polymerase that uses its integral RNA as a template to synthesize telomeric repeats onto chromosome ends. The molecular mechanism of telomerase is unique and involves a translocation step after the synthesis of each telomeric repeat. To directly measure the enzym...
Trapping of proteins under physiological conditions in a nanopipette.
Angewandte Chemie (International Edition) 44(24):3747 (2005)
Application of computational mechanics to the analysis of natural data: an example in geomagnetism.
Physical Review E 67(1 Pt 2):016203 (2003)
We discuss how the ideal formalism of computational mechanics can be adapted to apply to a noninfinite series of corrupted and correlated data, that is typical of most observed natural time series. Specifically, a simple filter that removes the corruption that creates rare unphysical causal stat...