Unified Theory for Gel Electrophoresis and Gel Filtration

Abstract

Abstract. Unified theory for gel electrophoresis and gel filtration: The behavior of macromolecules in gel filtration and gel electrophoresis may be predicted from Ogston's model for a random meshwork of fibers. This model has been generalized to apply to nonspherical molecules and to several gel types. The model provides equations for inter-relationships between mobility, partition coefficient, gel concentration, and molecular radius; it gives a non-Gaussian distribution of pore sizes as a function of gel concentration. The theory defines conditions for optimal separation and optimal resolution in gel filtration and gel electrophoresis. The difference in resolving power between the two fractionation methods is accounted for by the fact that gel filtration is a form of partition chromatography.Several models have been proposed for the mechanism of gel electrophoresis and gel filtration using diverse and mostly improbable assumptions regarding pore geometry.'-'0 The Ogston model makes only minimal assumptions regarding pore architecture, and is realistic in terms of known polymer structures such as polyacrylamide, cross-linked dextran, starch, agarose, or hyaluronic acid." 2 Ogston derived the distribution of spaces (fractional volume, f) available to a spherical molecule in a random suspension of linear fibers' or beads." 2 This model has been applied to gel filtration by making the assumption that the coefficient Ka. is equal tof;2 11where Ve is elution volume, Vo is void volume, and VT is total volume. Morris applied the Ogston model to gel electrophoresis"2 by proposing that the electrophoretic mobility (M) relative to free mobility (Mo) is equal tof:Morris noted that this provides a unified theoretical basis for gel electrophoresis and gel filtration, and suggested a linear relationship of the form:

Structural Characterization of PTX3 Disulfide Bond Network and Its Multimeric Status in Cumulus Matrix Organization

“…From the overall analysis of our MS data, we concluded that also Cys 47 establishes interchain symmetric disulfide bridges (see above). Site-directed mutagenesis results shown here are consistent with this and furthermore suggest that Cys 47 -Cy 47 and Cys 49 -Cys 49 disulfide bonds occur within the same pairs of PTX3 subunits. As can be seen from Fig.…”

“…Finally the signal at 210 kDa appeared more prominent as compared with that described for wt PTX3. The close similarity of the band patterns for C47S and C49S to that of the wt PTX3 indicates that Cys 47 and Cys 49 exert a redundant role in PTX3 covalent oligomerization. Cys 49 was shown by MS to form interchain symmetric disulfide bonds.…”

“…PTX3 encodes nine cysteines: three in the N-terminal region and six in the C-terminal pentraxin domain (23). Fragments were detected containing every cysteine apart from Cys 47 probably because peptides containing Cys 47 have low ionization efficiency and/or low solubility under the LC conditions exploited. No peptide was detected covering Asn 220 when endoproteinase incubations were performed on glycosylated PTX3.…”

“…In contrast to CRP and SAP, PTX3 oligomers are stabilized by covalent bonds (23). The mature protein contains nine cysteine residues: three are located within the N-terminal region (Cys 47 , Cys 49 , and Cys 103 ), and six are in the C-terminal domain (Cys 179 , Cys 210 , Cys 271 , Cys 317 , Cys 318 , and Cys 357 ). Cys residues at positions 210 and 271 are highly conserved among pentraxins and based on the homology with CRP and SAP are predicted to be engaged in an intrachain disulfide bond (22,23).…”

Evaluation of capillaries with different inner coatings for DNA analysis using dilute polymer solutions by capillary electrophoresis

“…The Ogston mechanism describes satisfactorily the mobility of small DNA fragments in low electric fields and in low gel concentrations. [8][9][10][11][12] When a larger DNA fragment crosses a polymeric network with ξ << Rg, it undergoes a distortion so that it still passes across the dynamic pore; therefore, there is no further agreement with the Ogston theory. The reptation system explains this fact, since it describes the DNA as a molecule with an undisturbed spiral conformation, which moves like a snake, "crawling" along the "tubes" formed by the polymeric matrix.…”