scholarly journals Challenges in estimating the motility parametersn of single processive motor proteins

2017 ◽  
Author(s):  
F. Ruhnow ◽  
L. Kloß ◽  
S. Diez
Keyword(s):  
Experimental Data ◽  
Single Molecule ◽  
Motor Proteins ◽  
Evaluation Method ◽  
Length Distribution ◽  
Acquisition Time ◽  
Filament Length ◽  
Experimental Conditions ◽  
Run Length ◽  
Wide Range

AbstractCytoskeletal motor proteins are essential to the function of a wide range of intracellular mechanosystems. The biophysical characterization of their movement along their filamentous tracks is therefore of large importance. Towards this end, single-molecule, in vitro stepping-motility assays are commonly used to determine motor velocity and run length. However, comparing results from such experiments has proved difficult due to influences from variations in the experimental conditions and the data analysis methods. Here, we investigate the movement of fluorescently-labeled, processive, dimeric motor proteins and propose a unified algorithm to correct the measurements for finite filament length as well as photobleaching. Particular emphasis is put on estimating the statistical errors associated with the proposed evaluation method as knowledge of these values is crucial when comparing measurements from different experiments. Testing our approach with simulated and experimental data from GFP-labeled kinesin-1 motors stepping along immobilized microtubules, we show (i) that velocity distributions should be fitted by a t location-scale probability density function rather than by a norm*al distribution, (ii) that the impossibility to measure events shorter than the image acquisition time needs to be accounted for, (iii) that the interaction time and run length of the motors can be estimated independent of the filament length distribution, and (iv) that the dimeric nature of the motors needs to be considered when correcting for photobleaching. Moreover, our analysis reveals that controlling the temperature during the experiments with a precision below 1 K is of importance. We believe, our method will not only improve the evaluation of experimental data, but will also allow for better statistical comparisons between different populations of motor proteins (e.g. with distinct mutations or linked to different cargos) and filaments (e.g. in distinct nucleotide states or with different posttranslational modifications).

Improved Measurement of the Kinetics of Crystallization in a Batch Experiment

1993 ◽  
Vol 58 (8) ◽  
pp. 1839-1847 ◽  
Author(s):  
Jaroslav Nývlt ◽  
Miloslav Karel
Keyword(s):  
Experimental Data ◽  
Crystal Size ◽  
Evaluation Method ◽  
Potassium Sulfate ◽  
Batch Experiment ◽  
Improved Method ◽  
Short Intervals ◽  
Wide Range ◽  
Kinetics Of ◽  
Single Batch

An improved method is described whereby knowledge of the supersaturation course during a run and measurement of the final product crystal size distribution yield the growth rate of crystals and the nucleation rate in a wide range of supersaturations from only a single batch experiment. The evaluation method is refined by the interpolation of experimental data for short intervals of time. The method is illustrated using potassium sulfate crystallization as an example.

scholarly journals Photochemical laws and visual phenomena

1946 ◽  
Vol 133 (870) ◽  
pp. 63-75 ◽  
Keyword(s):  
Experimental Data ◽  
Recovery Process ◽  
Visual Sensitivity ◽  
Rapid Recovery ◽  
Foveal Vision ◽  
Experimental Conditions ◽  
Reciprocity Law ◽  
Wide Range ◽  
Visual Phenomena ◽  
Nervous Transmission

By measuring the course of recovery of visual sensitivity in the dark, after exposure to light, it has been demonstrated that all the phenomena of recovery can be explained—qualitatively—by the known fundamental principles of photochemistry. In particular, the reciprocity law, which is widely valid in photographic processes, only holds under certain limited circumstances for visual phenomena. In the case of foveal vision the reciprocity law does not hold, owing to the rapid recovery process, except for very brief periods of exposure. In the case of parafoveal vision the recovery process for the scotopic mechanism is sufficiently slow to allow validity of the reciprocity law over a wide range of exposure periods, although the law still breaks down for the parafoveal photopic mechanism. This circumscribed validity of the reciprocity law is of practical service in that the effect of many experimental conditions can be predicted from a very limited set of suitably chosen experimental data. It may be noted that it has not been found necessary to introduce any postulate involving recovery of the nervous transmission system as part of the recovery process.

scholarly journals Systematic assessment of the accuracy of subunit counting in biomolecular complexes using automated single molecule brightness analysis

2021 ◽  
Author(s):  
John S H Danial ◽  
Yuri Quintana ◽  
Uris Ros ◽  
Raed Shalaby ◽  
Eleonora Germana Margheritis ◽  
...  
Keyword(s):  
Single Molecule ◽  
Ground Truth ◽  
Experimental Conditions ◽  
Systematic Analysis ◽  
Ground Truth Data ◽  
Systematic Assessment ◽  
Brightness Analysis ◽  
Wide Range ◽  
Biomolecular Complexes ◽  
Biological Complexes

Analysis of single molecule brightness allows subunit counting of high-order oligomeric biomolecular complexes. Although the theory behind the method has been extensively assessed, systematic analysis of the experimental conditions required to accurately quantify the stoichiometry of biological complexes remains challenging. In this work, we develop a high-throughput, automated computational pipeline for single molecule brightness analysis that requires minimal human input. We use this strategy to systematically quantify the accuracy of counting under a wide range of experimental conditions in simulated ground-truth data and then validate its use on experimentally obtained data. Our approach defines a set of conditions under which subunit counting by brightness analysis is designed to work optimally and helps establishing the experimental limits in quantifying the number of subunits in a complex of interest. Finally, we combine these features into a powerful, yet simple, software that can be easily used for the stoichiometry analysis of such complexes.

scholarly journals Growth analysis of Scots pine (Pinus sylvestris L.) seedlings cultivated in a wide range of experimental conditions

2014 ◽  
Vol 54 (3) ◽  
pp. 255-272 ◽  
Author(s):  
Włodzimierz Żelawski ◽  
Andrzej Łotocki ◽  
Helena Morteczka ◽  
Teresa Przykorska-Żelawska ◽  
Wincenty Wrrześniewski
Keyword(s):  
Experimental Data ◽  
Plant Growth ◽  
Scots Pine ◽  
Growth Analysis ◽  
Time Instant ◽  
Initial Value ◽  
Experimental Conditions ◽  
Pine Seedlings ◽  
Wide Range ◽  
Future Work

The work of this laboratory for many years with Scots pine seedlings has allowed to collect quite unique information as to the accumulation and distribution of dry matter and the reaction of plants to various environmental influences. It seems worthwhile to publish the main experimental data almost <em>in extenso</em> because they may be used in future work for building mathematical models of plant growth. The presented experimental data enabled to discuss controversial points of quantitative analysis of plant growth such as: the time instant and the initial value of growth to be taken as the starting points (t<sub>0</sub> W<sub>0</sub>); exponential approximation of the growth curve; inadequacy of allometric approach in modelling vegetative growth of plants, etc. The discussion aims at defining more closely the essential notions of growth analysis.

scholarly journals Drag-induced directionality switching of kinesin-5 Cin8 revealed by cluster-motility analysis

2020 ◽  
Author(s):  
Himanshu Pandey ◽  
Emanuel Reithmann ◽  
Alina Goldstein-Levitin ◽  
Jawdat Al-Bassam ◽  
Erwin Frey ◽  
...  
Keyword(s):  
Single Molecule ◽  
Motor Proteins ◽  
Molecular Model ◽  
Mitotic Cell ◽  
Underlying Mechanism ◽  
General Mechanism ◽  
Experimental Conditions ◽  
Active Motion ◽  
Asymmetric Response ◽  
External Forces

AbstractDirected active motion of motor proteins is a vital process in virtually all eukaryotic cells. Nearly a decade ago, the discovery of directionality switching of mitotic kinesin-5 motors challenged the long-standing paradigm that individual kinesin motors are characterized by an intrinsic directionality. While several kinesin motors have now been shown to exhibit context-dependent directionality that can be altered under diverse experimental conditions, the underlying mechanism remains unknown. Here, we studied clustering-induced directionality switching of the mitotic kinesin-5 Cin8, using a fluorescence-based single-molecule motility assay combined with biophysical theory. Based on the detailed characterization of the motility of single motors and clusters of Cin8, we developed a predictive molecular model, that quantitatively agrees with experimental data. This combined approach allowed us to quantify the response of Cin8 motors to external forces as well as the interactions between Cin8 motors, and thereby develop a detailed understanding of the molecular mechanism underlying directionality switching. The main insight is that directionality switching is caused by a single feature of Cin8: an asymmetric response of active motion to forces that oppose motion, here referred to as drag. This general mechanism explains why bidirectional motor proteins are capable of reversing direction in response to seemingly unrelated experimental factors including clustering, changes in the ionic strength of the buffer, increased motor density and molecular crowding, and in motility assays.Significance StatementKinesin-5 motor proteins perform essential functions in chromosome segregation during mitotic cell division. Surprisingly, several kinesin-5 motors have the ability to reverse directionality under different experimental conditions, which contradicts the long-standing paradigm that individual kinesin motors are characterized by an intrinsic directionality. The mechanism underlying this ability to switch directionality has remained elusive. Here, we combine fluorescence-based motility assays and theoretical modeling to analyze cluster-size-dependent motility of the bidirectional kinesin-5 Cin8. Our results show that bidirectional motors can switch directionality because they exhibit an asymmetric response of active motion to drag. This mechanism explains multiple seemingly unrelated experimental factors that have been shown to cause directionality switching of kinesin motors.

scholarly journals Comparing thixotropic and Herschel–Bulkley parameterizations for continuum models of avalanches and subaqueous debris flows

2018 ◽  
Vol 18 (1) ◽  
pp. 303-319 ◽  
Author(s):  
Chan-Hoo Jeon ◽  
Ben R. Hodges
Keyword(s):  
Experimental Data ◽  
Debris Flows ◽  
Fluid Model ◽  
Liquid Mixtures ◽  
High Yield ◽  
Aging Effects ◽  
Stress Strain ◽  
Experimental Conditions ◽  
Wide Range ◽  
Strain Relationship

Abstract. Avalanches and subaqueous debris flows are two cases of a wide range of natural hazards that have been previously modeled with non-Newtonian fluid mechanics approximating the interplay of forces associated with gravity flows of granular and solid–liquid mixtures. The complex behaviors of such flows at unsteady flow initiation (i.e., destruction of structural jamming) and flow stalling (restructuralization) imply that the representative viscosity–stress relationships should include hysteresis: there is no reason to expect the timescale of microstructure destruction is the same as the timescale of restructuralization. The non-Newtonian Herschel–Bulkley relationship that has been previously used in such models implies complete reversibility of the stress–strain relationship and thus cannot correctly represent unsteady phases. In contrast, a thixotropic non-Newtonian model allows representation of initial structural jamming and aging effects that provide hysteresis in the stress–strain relationship. In this study, a thixotropic model and a Herschel–Bulkley model are compared to each other and to prior laboratory experiments that are representative of an avalanche and a subaqueous debris flow. A numerical solver using a multi-material level-set method is applied to track multiple interfaces simultaneously in the simulations. The numerical results are validated with analytical solutions and available experimental data using parameters selected based on the experimental setup and without post hoc calibration. The thixotropic (time-dependent) fluid model shows reasonable agreement with all the experimental data. For most of the experimental conditions, the Herschel–Bulkley (time-independent) model results were similar to the thixotropic model, a critical exception being conditions with a high yield stress where the Herschel–Bulkley model did not initiate flow. These results indicate that the thixotropic relationship is promising for modeling unsteady phases of debris flows and avalanches, but there is a need for better understanding of the correct material parameters and parameters for the initial structural jamming and characteristic time of aging, which requires more detailed experimental data than presently available.

Smoothing of a stationary random signal in a continuous mixer

1987 ◽  
Vol 52 (7) ◽  
pp. 1730-1741
Author(s):  
Pavel Hasal ◽  
Jitka Vyhlídková ◽  
Vladimír Kudrna
Keyword(s):  
Experimental Data ◽  
Residence Time ◽  
Theoretical Concept ◽  
Random Signal ◽  
Smoothing Effect ◽  
Experimental Conditions ◽  
Stochastic Fluctuations ◽  
Wide Range ◽  
Concentration Signal ◽  
Good Agreement

The paper presents results of the first stage of experimental verification of the theoretical concept published in the two first preceding communications concerning the flow of liquid in a continuous nonideal mixer and, namely, the smoothing effect of such a mixer on a passing stationary random concentration signal. In a sufficiently wide range of experimental conditions it has been proven that stochastic fluctuations of the distribution of the residence time of liquid in the mixer do not contribute significantly to the variance of the outlet random signal of the mixer. Further, a good agreement has been proven between the experimental data and the model relationship for the smoothing effect of the mixer based on the application of the gamma distribution of the residence time.

Microchannel Condensation: Comparison of Annular Laminar Flow Theory With Detailed Measurements

2013 ◽  
Author(s):  
H. S. Wang ◽  
J. W. Rose
Keyword(s):  
Experimental Data ◽  
Heat Flux ◽  
Laminar Flow ◽  
Local Heat ◽  
Experimental Investigations ◽  
Experimental Conditions ◽  
Two Fluids ◽  
Wide Range ◽  
Local Heat Flux ◽  
Good Agreement

Detailed experimental investigations of condensation in microchannels where local heat flux and surface temperature were measured along the channel are compared with theoretical results for the special case of annular, laminar flow. The theoretical model includes surface tension driven transverse flow towards the corners of the channel as well as shear stress driven streamwise flow in an otherwise Nusselt treatment. The theory has no empirical input. When distributions along the channel of the local vapor and wall temperatures are given, local heat flux and heat-transfer coefficient, as well as local vapor quality, may be calculated. Such detailed experimental data have only recently become available. Strict implementation of the theory requires that the onset of condensation occurs within the channel, i.e. the vapor is saturated or superheated at the inlet. The comparisons show remarkably good agreement with the experimental data for two fluids and covering a wide range of experimental conditions.

An Improved Generalized Model for Predicting Frost Growth on a Cold Flat Plate

2010 ◽  
Author(s):  
Q. C. Guo ◽  
W. Wang ◽  
J. Xiao ◽  
W. P. Lu
Keyword(s):  
Experimental Data ◽  
Flat Plate ◽  
Current Model ◽  
Theoretical Models ◽  
Maximum Error ◽  
Open Loop ◽  
Experimental Conditions ◽  
Wide Range ◽  
Semi Empirical ◽  
Frost Growth

A generalized quasi-steady and one-dimensional model for predicting the frost growth on flat plate was proposed based on the previous theoretical models. To improve the predicting ability of the current model, a modified semi-empirical correlation for calculating initial condition of frost density was presented experimentally. The experiments were conducted in a suction-type open-loop wind tunnel under a series of experimental conditions: air temperature −8°C to 19°C, humidity 42% to 80%, velocity 5m/s and the temperature of cold plate −16°C to −8°C. The numerical results of frost thickness, frost density, frost surface temperature and heat flux rate were compared to the experimental data. The simulation results were found agree with the experimental results in a maximum error of 10%. The presented model was further validated by comparing with the previous published experimental data in a wide range of frosting conditions. It was found that the presented model was a simple but universal one to predict the frost growth on cold flat plate.

scholarly journals An Automated Bayesian Pipeline for Rapid Analysis of Single-Molecule Binding Data

2018 ◽  
Author(s):  
Carlas S. Smith ◽  
Karina Jouravleva ◽  
Maximiliaan Huisman ◽  
Samson M. Jolly ◽  
Phillip D. Zamore ◽  
...  
Keyword(s):  
Single Molecule ◽  
Thermus Thermophilus ◽  
Binding Assays ◽  
Binding Properties ◽  
Experimental Conditions ◽  
High Throughput Analysis ◽  
Binding Data ◽  
Wide Range ◽  
Automated Pipeline ◽  
And Function

SummarySingle-molecule binding assays enable the study of how molecular machines assemble and function. Current algorithms can identify and locate individual molecules, but require tedious manual validation of each spot. Moreover, no solution for high-throughput analysis of single-molecule binding data exists. Here, we describe an automated pipeline to analyze single-molecule data over a wide range of experimental conditions. We benchmarked the pipeline by measuring the binding properties of the well-studied, DNA-guided DNA endonuclease, TtAgo, an Argonaute protein from the Eubacterium Thermus thermophilus. We also used the pipeline to extend our understanding of TtAgo by measuring the protein’s binding kinetics at physiological temperatures and for target DNAs containing multiple, adjacent binding sites.

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