Disc gel electrophoresis,

Hedrick, JL Smith, AJ, Size and Charge Isomer Separation and Estimation of Molecular Weights of Proteins by Disc Gel Electrophoresis, Archives of Biochemistry and Biophysics 126, 155, 1968. 

Chromatography and Electrophoresis of Antigens. Cotton dust (20-30 mg) was dissolved in water and passed through a small column (30 x 1.5 cm id) packed with 3 g of Sephadex G-75 that had been preswelled in water. Chromatograms were obtained with an LKB Uvicord ultraviolet monitor and a chopper bar recorder. Volumes of 5 ml each were collected in test tubes with an automatic fraction collector. Two components were detected at 250 nm. Samples of 10 mg were analyzed by disc gel electrophoresis according to procedures described by Davis (36), and Zacharius (37). 

The process of disc gel electrophoresis. A Before electrophoresis. B Movement of chloride, glycinate, and protein through the stacking gel. 

Disc gel electrophoresis yields excellent resolution and is the method of choice for analysis of proteins and nucleic acid fragments. Protein or nucleic acid bands containing as little as 1 or 2 ju,g can be detected by staining the gels after electrophoresis. 

Purified preparations of alkaline phosphatase from E. coli, judged homogeneous when examined in the analytical ultracentrifuge, contain several isozymes, because several bands which contain enzymic activity are obtained in starch-gel and disc-gel electrophoresis. Although most workers find three bands (38, 39, 41, 43, 69), four (44) and five (70) equally spaced bands have been found. 

Lazdunski and Lazdunski (43), separated three isozymes on DEAE-cellulose. They found that pure samples of either isozyme I (the isozyme with the least negative charge at pH 7.0 is referred to as isozyme I) or isozyme III, after dissociation and reassociation, gave only the original single band on disc-gel electrophoresis. Pure isozyme II, after dissociation and reassociation, gave three bands, corresponding to isozymes I, II, and III. It was later shown (44), that when monomers of isozymes I and III are mixed before reassociation, three bands are obtained. This... 

The cellulase components that are synthesized in the presence of sophorose were investigated by the basic procedures previously described (1,2,4) for the isolation of cellulolytic components from commercial cellulase preparations. The purification to homogeneity of the proteins that yield the three predominant bands when the crude preparation is subjected to disc gel electrophoresis was accomplished by ion exchange chromatography. 

Figure 9. Polyacrylamide disc gel electrophoresis of soluble SOD from (A) Spinacia (spinach) (a, b), Spirulina (blue-green alga) (c,d), ana from (B) rod outer segments from retina of cattle (a,b,d) and frogs (c,e) (39, 64)...

Myosin. Rabbit muscle myosin is a long, thin molecule (VI400 X 20-50 A) with a molecular weight of 5 X 10. It is composed of two heavy chains and four light chains as demonstrated by SDS-polyacrylamide disc gel electrophoresis. On tryptic digestion, myosin is split into the subunits, H-meromyosin (HMM) and L-mero-myosin (LMM). HMM is further split into S-l and S-2 subunits. While LMM is a rod of V)0% a-helical content, the a-helical content for HMM, S-l and S-2 fragments is 46%, 33% and 87%, respectively. The ATPase activity is localized in the S-l subunit (33,34). Although fish myosins appear to have the same structural profile (10,22,35-40) and similar amino acid composition as rabbit myosin (39,41,42), fish myosin is different from rabbit myosin in physicochemical properties such as solubility, viscosity and stability (10,22,35-40). 

However, our work on in vitro frozen storage of isolated carp actomyosin showed that actin is denatured progressively with myosin as demonstrated by SDS-polyacrylamide disc gel electrophoresis (90). 

Etzler and Kabat purified the agglutinating principle from Dolichos seed extracts by adsorption to insoluble polyleucyl hog A + H substance.108,517 Specific elution of the adsorbent with 2-acetamido-2-deoxy-D-galactose gave a protein that appeared homogeneous by immunodiffusion, immunoelectrophoresis, disc-gel electrophoresis under acid and alkaline conditions, and sedimentation analysis. The amino acid composition of the protein of molecular weight 141,000 reflected a high content of aspartic acid and serine, little methionine, and no cysteine. Carbohydrate analysis showed a sugar content of 2.4% of hexose, 1.6% of hexosamine, and 1.5% of 2-acetamido-2-deoxy-hexose. 

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