Electrophoresis sodium dodecyl sulfate

Fig. 2. Covalent attachment of poly(ADPR) to the nonhistone proteins being retained by boronate agarose chromatography. The fraction of nonhistone proteins being retained by the boronate resin was separated by sodium dodecyl sulfate electrophoresis (cf. Fig. 1). The stained gel was sliced and poly(ADPR) determined in individual fractions utilizing a specific radioimmunoassay [1]...

Fig. 4. Identification of ADP-ribosylated topoisomerase I following renaturation (cf. Fig. 3). The reaction mixture for the topoisomerase I assay contained in a total volume of 15 /ul 50 mAf Tris/ HCl, pH 7.5, 50 mM KCl, 10 mMMgCl, 0.5 mAf DTT, 0.1 mA/EDTA, 0.03 mg mr BSA, 0.5 Mg of form I phi X 174 DNA (BRL), and 10 m1 of renatured and diluted poly(ADPR) nonhistone proteins. ATP and spermidine were included where indicated. The incubation was carried out for 30 min at 37 C and was terminated by the addition of 2 til of proteinase K (Merck) (2 mg ml of 1% sodium dodecyl sulfate). Electrophoresis was performed for 15 h at 50 V using an 1.4% agarose gel and 1 Mg ml ethidium bromide for staining. Lane 1 control without enzyme lanes 2—4 8-, 16-, and 32-fold diluted samples lanes 5—7 8-, 16-, and 32-fold diluted samples incubated in the presence of 1 vaM ATP lanes 8-10 4-, 8-, and 16-fold diluted samples incubated in the presence of 1 mM ATP and 5 mM spermidine...

Microtubule-associated proteins bind to microtubules in vivo and subserve a number of functions including the promotion of microtubule assembly and bundling, chemomechanical force generation, and the attachment of microtubules to transport vesicles and organelles (Olmsted, 1986). Tubulin purified from brain tissue by repeated polymerization-depolymerization contains up to 20% MAPs. The latter can be dissociated from tubulin by ion-exchange chromatography. The MAPs from brain can be resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). 

The protein was purified by a dialysis procedure, denatured and analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Western blotting indicated that the protein of interest consisted of two components, one of which increased in concentration as the purification proceeded. The authors initially suggested that this could be due to the presence of a number of species produced by modification of the amino acid side-chains, for example, by glyco-sylation, or by modification of the C- or N- terminus. 

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) An electrophoretic technique used for the separation of proteins. 

Guttman, A, On the Separation Mechanism of Capillary Sodium Dodecyl Sulfate-Gel Electrophoresis of Proteins, Electrophoresis 16, 611, 1995. 

The number of different proteins in a membrane varies from less than a dozen in the sarcoplasmic reticulum to over 100 in the plasma membrane. Most membrane proteins can be separated from one another using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), a technique that has revolutionized their study. In the absence of SDS, few membrane proteins would remain soluble during electrophoresis. Proteins are the major functional molecules of membranes and consist of enzymes, pumps and channels, structural components, antigens (eg, for histocompatibility), and receptors for various molecules. Because every membrane possesses a different complement of proteins, there is no such thing as a typical membrane structure. The enzymatic properties of several different membranes are shown in Table 41-2. 

Guttman, A., Shieh, P, Lindahl, J., and Cooke, N., Capillary sodium dodecyl sulfate gel electrophoresis of proteins. II. On the Ferguson method of polyethylene oxide gels, /. Chromatogr., 676, 227, 1994. 

Benedek, K. and Thiede, S., High performance capillary electrophoresis of proteins using sodium dodecyl sulfate-poly(ethylene oxide), J. Chromatogr., 676, 209, 1994. 

Guttman, A. and Nolan, J., Comparison of the separation of proteins by sodium dodecyl sulfate-slab gel electrophoresis and capillary sodium dodecyl sulfate-gel electrophoresis, Anal. Biochem., 221, 285, 1994. 

Dittman, M. M. and Rozing, G. R, High-sensitivity separations of sodium dodecyl sulfate-protein complexes with capillary gel electrophoresis, LC-GC, 17(2), 132, 1999. 

Hunt G. and Nashabeh W., Capillary electrophoresis sodium dodecyl sulfate nongel sieving analysis of a therapeutic recombinant monoclonal antibody a biotechnology perspective, Anal. Chem. 71, 2390,1999. 

A. Gerstner, Z. Csapo, M. Sasvari-Szekely, and A. Guttman, Ultrathin sodium dodecyl sulfate gel electrophoresis of proteins Effect of gel composition and temperature on the separation of sodium dodecyl sulfate-protein complexes, Electrophoresis, 21, 834 (2000). 

Prokaryotic cells express hundreds to thousands of proteins while higher eukaryotes express thousands to tens of thousands of proteins at any given time. If these proteins are to be individually identified and characterized, they must be efficiently fractionated. One-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) has typically been use to study protein mixtures of <100 proteins. Onedimensional electrophoresis is useful because nearly all proteins are soluble in SDS, molecules ranging from approximately 10,000 to 300,000 molecular weight can be resolved, and extremely basic or acidic proteins can be visualized. The major disadvantage to one-dimensional gels is that they are not suitable for complex mixtures such as proteins from whole cell lysates. 

Innovations in separation science continued on this theme and provided one of the most powerful separation techniques used in biochemistry, where proteins are separated with isoelectric focusing (IEF) applied in one direction, and gel electrophoresis (GE) applied at aright angle to the first separation direction (O Farrell, 1975 Celis and Bravo, 1984). In this case, proteins are first separated according to their isoelectric point, measured in p/units, and then according to their molecular weight by gel electrophoresis. The size separation step is usually aided by addition of a surfactant, most typically sodium dodecyl sulfate (SDS), and the gel material is a polyacrylamide formulation. 

Hayashi, T., Nagai, Y. (1980). The anomalous behavior of collagen peptides on sodium dodecyl sulfate-polyacrylamide gel electrophoresis is due to the low content of hydrophobic amino acid residues. J. Biochem. (Tokyo) 87, 803-808. 

Karim, M.R., Shinagawa, S., Takagi, T. (1994). Electrophoretic mobilities of the complexes between sodium dodecyl sulfate and various peptides or proteins determined by free solution electrophoresis using coated capillaries. Electrophoresis 15, 1141-1146. 

Takagi, T., Tsujii, K., Shirahama, K. (1975). Binding isotherms of sodium dodecyl sulfate to protein polypeptides with special reference to SDS-polyacrylamide gel electrophoresis. J. Biochem. (Tokyo) 77, 939-947. 

Lin et al. [95] used capillary electrophoresis with dual cyclodextrin systems for the enantiomer separation of miconazole. A cyclodextrin-modified micellar capillary electrophoretic method was developed using mixture of /i-cyclodextrins and mono-3-0-phenylcarbamoyl-/j-cyclodextrin as chiral additives for the chiral separation of miconazole with the dual cyclodextrins systems. The enantiomers were resolved using a running buffer of 50 mmol/L borate pH 9.5 containing 15 mmol/L jS-cyclodextrin and 15 mmol/L mono-3-<9-phcnylcarbamoyl-/j-cyclodextrin containing 50 mmol/L sodium dodecyl sulfate and 1 mol/L urea. A study of the respective influence of the /i-cyclodcxtrin and the mono-3-(9-phenylcarbamoyl-/i-cyclodextrin concentration was performed to determine the optical conditions with respect to the resolution. Good repeatability of the method was obtained. 

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