Computer simulation and experimental validation of the electrophoretic behavior of proteins

Mosher, Richard A.; Dewey, Douglas.; Thormann, Wolfgang; Saville, D. A.; Bier, M.

doi:10.1021/ac00179a015

Cited by 126 publications

(101 citation statements)

References 20 publications

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“…This is illustrated here with ZE, ITP, IEF and EKC separation examples. The data shown were produced with GENTRANS, the simulator based on the work of Bier, Mosher, Thormann, Breadmore and coworkers [1,37,38,44,[49][50][51][52][53][54]75].…”

Section: Simulation Examplesmentioning

confidence: 99%

“…In a series of efforts, Mosher et al [44,49,130] expanded GENTRANS to be applicable for proteins. As proteins are complicated molecules with a large number of ionizable groups the mobility changes significantly as a function of pH.…”

Section: Aspects Of Itp and Mbementioning

confidence: 99%

“…As proteins are complicated molecules with a large number of ionizable groups the mobility changes significantly as a function of pH. A table of pH titration data (pH versus net charge) and an equation describing the impact of ionic strength are used to calculate the mobility of the protein [44]. After their initial work demonstrating the applicability to proteins, two algorithms to account for the influence of ionic strength on protein mobility were evaluated [49] with the best system subsequently used to examine the feasibility of using titration data generated from the amino acid composition and free amino acid pK a values [130].…”

Section: Aspects Of Itp and Mbementioning

confidence: 99%

“…Distortion of protein zones found in the collected fractions of continuous flow ITP, which was not observed for low molecular mass compounds and was not observed in the simulations, was attributed to the high density/viscosity gradient formed at the buffer interfaces as well as the design of the outlet ports. Simulation was also employed (i) to find proper low molecular mass spacers that established themselves between ITP zones of proteins [49,130] and could thus be used to fractionate human serum by recycling ITP [156], and (ii) to predict ITP zone formation of proteins [44] and low molecular mass substances [157] in configurations used for continuous flow ITP.…”

Section: Aspects Of Itp and Mbementioning

confidence: 99%

“…New models with sophisticated integration schemes emerged and the number of applications increased. Models that are currently in use in various laboratories are the 1-D simulators GENTRANS of Mosher et al [44,49,[51][52][53][54][129][130][131][132] and SIMUL5 of Hruška et al [48] (available at http://www.natur.cuni.cz/ Gas), which succeeded SIMUL 4.0 [47]. Interesting new models are those of Bercovici et al [69] (available as the Stanford's Public Release Electrophoretic Separation Solver (SPRESSO), http://microfluidics.stanford.edu/spresso), the 2-D model of Shim et al [70,71,135,136] and the models of Yang and his collaborators [68,138].…”

Section: Introductionmentioning

confidence: 99%

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Dynamic computer simulations of electrophoresis: A versatile research and teaching tool

et al. 2010

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Software is available, which simulates all basic electrophoretic systems, including moving boundary electrophoresis, zone electrophoresis, ITP, IEF and EKC, and their combinations under almost exactly the same conditions used in the laboratory. These dynamic models are based upon equations derived from the transport concepts such as electromigration, diffusion, electroosmosis and imposed hydrodynamic buffer flow that are applied to user-specified initial distributions of analytes and electrolytes. They are able to predict the evolution of electrolyte systems together with associated properties such as pH and conductivity profiles and are as such the most versatile tool to explore the fundamentals of electrokinetic separations and analyses. In addition to revealing the detailed mechanisms of fundamental phenomena that occur in electrophoretic separations, dynamic simulations are useful for educational purposes. This review includes a list of current high-resolution simulators, information on how a simulation is performed, simulation examples for zone electrophoresis, ITP, IEF and EKC and a comprehensive discussion of the applications and achievements.

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Section: Simulation Examplesmentioning

confidence: 99%

Section: Aspects Of Itp and Mbementioning

confidence: 99%

Section: Aspects Of Itp and Mbementioning

confidence: 99%

Section: Aspects Of Itp and Mbementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Dynamic computer simulations of electrophoresis: A versatile research and teaching tool

et al. 2010

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High-resolution computer simulation of the dynamics of isoelectric focusing using carrier ampholytes: The post-separation stabilizing phase revisited

Mosher¹,

Thormann

2002

Electrophoresis

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A dynamic electrophoresis simulator that accepts 150 components and voltage gradients employed in the laboratory was used to provide a detailed description of the stabilizing phase in isoelectric focusing under conditions that were hitherto inaccessible. High-resolution focusing data are presented for pH gradients spanning 7 units (pH 3-10 and pH 4-11 with 20 carrier ampholytes/pH unit) and 3.5 units (pH 7-10.5 and pH 5-8.5 with 40 carrier ampholytes/pH unit). Stabilizing phase behavior for configurations (i) with the focusing column ends only permeable to OH(-) and H(+) at cathode and anode, respectively, and (ii) with the focusing column being sandwiched between NaOH (catholyte) and phosphoric acid (anolyte) are described. Simulation data reveal the stabilizing phase to be diffusion-controlled and characterized by changes that progress from the column ends towards neutrality (i.e., towards the center in case of pH gradients bracketing neutrality). Transient states are characterized by moving concentration valleys of carrier ampholytes that significantly alter the distributions of pH and conductivity. Nonlinear pH gradients are produced. The magnitude of the changes occuring is dependent on the span of the pH gradient. Gradients that encompass greater extremes of pH show more pronounced stabilizing phases. For all systems subjected to a constant 300 V/cm, the initial separation and subsequent stabilization require less than 10 min and more than 7000 min, respectively. The presence of electrolytes at the column ends disrupts the stabilizing phase, with the degree of disruption dependent on the concentrations of the acid and base employed as electrode solutions. The data not only indicate that a true steady state is never attained in the average laboratory experiment, they also suggest that a true steady state in absence of immobilized pH gradients cannot be achieved experimentally at all.

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Up-scaling capillary zone electrophoresis separations of polydisperse anionic polyelectrolytes with preparative free-flow electrophoresis exemplified with a soil fulvic acid

Junkers¹,

Schmitt‐Kopplin

Munch³

et al. 2002

Electrophoresis

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A scale-up of analytical capillary zone electrophoresis (CZE) to preparative free-flow electrophoresis (FFE) is described. FFE allows fractionations based on charge densities in larger amounts than in CZE, enabling further off-line analysis of the fractions. Model compounds (carboxylic acids and polystyrene sulfonates) showed a similar behavior in FFE as in CZE. Diffusion and electrodynamic distortion effects are more pronounced in FFE than in CZE. A soil fulvic acid was analyzed by CZE and fractionated by FFE. A comparison of the FFE fractions with CZE measurements of the same sample using the effective mobility scale showed good agreement of the two methods.

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Computer simulation and experimental validation of the electrophoretic behavior of proteins

Cited by 126 publications

References 20 publications

Dynamic computer simulations of electrophoresis: A versatile research and teaching tool

Dynamic computer simulations of electrophoresis: A versatile research and teaching tool

High-resolution computer simulation of the dynamics of isoelectric focusing using carrier ampholytes: The post-separation stabilizing phase revisited

Up-scaling capillary zone electrophoresis separations of polydisperse anionic polyelectrolytes with preparative free-flow electrophoresis exemplified with a soil fulvic acid

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