Proteins dialysis

Remove excess crosslinker and reaction by-products by dialysis or size exclusion chromatography. For small quantities of bait proteins, dialysis may be the better choice, because gel filtration columns often bind nonspecifically enough protein to make recoveries unacceptably low. 

Figure 2.5 As with all the other methods a great many varieties exist for executing the dialysis method. For not too small amounts of protein, dialysis tubes as shown in the figure can be used. The dialysis membrane is attached to the tube by a rubber ring. Figure kindly supplied by J. Drenth, University of Groningen and reproduced with permission.

Persons using dialysis require restriction of sodium, potassium and water intake. Protein intake is also closely controlled and will vary from 30 to 60 g daily. The aim is to provide about 3/4 of the protein allowance as protein of high biological value. Furthermore, to prevent high blood urea and potassium levels, it is especially important that adequate energy be supplied to prevent the catabolism of body protein. Dialysis patients also require vitamin supplementation in addition to the dietary controls. 

Films or membranes of silkworm silk have been produced by air-drying aqueous solutions prepared from the concentrated salts, followed by dialysis (11,28). The films, which are water soluble, generally contain silk in the silk I conformation with a significant content of random coil. Many different treatments have been used to modify these films to decrease their water solubiUty by converting silk I to silk II in a process found usehil for enzyme entrapment (28). Silk membranes have also been cast from fibroin solutions and characterized for permeation properties. Oxygen and water vapor transmission rates were dependent on the exposure conditions to methanol to faciUtate the conversion to silk II (29). Thin monolayer films have been formed from solubilized silkworm silk using Langmuir techniques to faciUtate stmctural characterization of the protein (30). ResolubiLized silkworm cocoon silk has been spun into fibers (31), as have recombinant silkworm silks (32). 

Flavin mononucleotide was first isolated from the yellow en2yme in yeast by Warburg and Christian in 1932 (4). The yellow en2yme was spHt into the protein and the yellow prosthetic group (coen2yme) by dialysis under acidic conditions. Flavin mononucleotide was isolated as its crystalline calcium salt and shown to be riboflavin-5Lphosphate its stmeture was confirmed by chemical synthesis by Kuhn and Rudy (94). It is commercially available as the monosodium salt dihydrate [6184-17 /, with a water solubiUty of more than 200 times that of riboflavin. It has wide appHcation in multivitamin and B-complex solutions, where it does not require the solubili2ers needed for riboflavin. 

Urea Enzymatic Dialysis Method. This method (16) uses 8 M urea [57-13-6] to gelatinize and facUitate removal of starch and promote extraction of the soluble fiber at mild (50°C) temperatures. EoUowing digestion with heat-stable a-amylase and protease, IDE is isolated by filtration or I DE is obtained after ethanol precipitation. Values for I DE are comparable to those obtained by the methods described eadier, and this method is less time-consuming than are the two AO AC-approved methods. Corrections for protein are required as in the AO AC methods. 

Lectins (proteins and/or glycoproteins of non-immune origin that agglutinate cells, from seeds of Robinia pseudoacacia), M 100,000. Purified by pptn with ammonium sulfate and dialysis then chromatographed on DE-52 DEAE-cellulose anion-exchanger, hydroxylapatite and Sephacryl S-200. [Wantyghem et al. Biochem J 237 483 1986.]... 

If a solution of protein is separated from a bathing solution by a semipermeable membrane, small molecules and ions can pass through the semipermeable membrane to equilibrate between the protein solution and the bathing solution, called the dialysis bath or dialysate (Figure 5A.2). This method is useful for removing small molecules from macromolecular solutions or for altering the composition of the protein-containing solution. 

Ultrafiltration is an improvement on the dialysis principle. Filters having pore sizes over the range of biomolecular dimensions are used to filter solutions to select for molecules in a particular size range. Because the pore sizes in these filters are microscopic, high pressures are often required to force the solution through the filter. This technique is useful for concentrating dilute solutions of macromolecules. The concentrated protein can then be diluted into the solution of choice. 

Ribosomal protein L12 was oxidized with 0.3 M H202 at 30°C for 1 h. After dialysis, the protein was incubated in the presence of 0.8 M 2-mercaptoethanol for 48 min at 37 °C and dialyzed. The amount of methionine residues was quantitated by exhaustive alkylation of the protein with [14C]iodoacetic acid. 

Water soluble protein with a relative molecular mass of ca. 32600, which particularly contains copper and zinc bound like chelate (ca. 4 gram atoms) and has superoxide-dismutase-activity. It is isolated from bovine liver or from hemolyzed, plasma free erythrocytes obtained from bovine blood. Purification by manyfold fractionated precipitation and solvolyse methods and definitive separation of the residual foreign proteins by denaturizing heating of the orgotein concentrate in buffer solution to ca. 65-70 C and gel filtration and/or dialysis. 

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. 

Erythropoietin 166 amino acids glycosylated Mammalian cells Treatment of anaemia associated with dialysis and AZT/AIDS Approved for sale Without glycosylation protein is cleared very quickly from plasma... 

Cultures from different times of growth were collected. Culture fluids were cleared by passing through glass fibre filter. After dialysis for 16-18 h against distilled water at 5°C, filtrates were assayed for enzyme activities and proteins. 

The specimens were analyzed at spectrophotometer at 280 nm. Preparations were purified from salts by dialysis. Protein concentrations in the initial and purified enzyme preparations were determined by Lowry method [6]. 

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