Protein fractionation chromatography

Although the total content of carbohydrate fractions of the three components is similar, as reported by Williams et al., 1990, it was found that protein-rich fractions have a significantly lower glucuronic acid content. Circular dichroism studies conducted on different GA fractions showed that only the AGP and GP components have a secondary structure (Renard et al., 2006). The AGP fraction was isolated by gel filtration chromatography and subjected to deglycosylation with hydrofluoric acid (HF) to separate the protein (Qi et al., 1991). About 400 amino acids were contained by the AGP protein fraction ( 33% are... 

Porath, J., Carlsson, J., Olsson, I., and Belfrage, G., Metal chelate affinity chromatography, a new approach to protein fractionation, Nature, 258, 598, 1975. 

Wienkoop, S., Glinski, M., Tanaka, N., Tolstikov, V.V., Fiehn, O., Weckwerth, W. (2004). Linking protein fractionation with multidimensional monolithic reversed-phase peptide chromatography/mass spectrometry enhances protein identification from complex mixtures even in the presence of abundant proteins. Rapid Commun. Mass Spectrom. 18, 643-650. 

Size-exclusion chromatography, also termed gel-permeation or gel-filtration chromatography, separates proteins on the basis of their size and shape. As most proteins fractionated by this technique are considered to have approximately similar molecular shape, separation is often described as being on the basis of molecular mass, although such a description is somewhat simplistic. 

Hydroxyapatite occurs naturally as a mineral in phosphate rock and also constitutes the mineral portion of bone. It may also be used to fractionate protein by chromatography. 

Figure 3 Biosynthesis and purification of 90-kD elastin analogue analyzed by denaturing polyacrylamide gel electrophoresis (10-15% gradient, visualized by silver staining). Lanes 1-7 time course of target protein expression at 0, 30, 60, 90, 120, 150, and 180 minutes after induction. Lane 9 soluble lysate of induced E. coli expression strain BLR(DE3)pRAMl. Lanes 10-13 protein fractions obtained from immobilized metal affinity chromatography of the lysate on nickel-NTA agarose (imidazole gradient elution). Lanes 8,14 protein molecular weight standards of 50, 75, 100, and 150 kD.

HPLC analysis of food proteins and peptides can be performed for different purposes to characterize food, to detect frauds, to assess the severity of thermal treatments, etc. To detect and/or quantify protein and peptide components in foods, a number of different analytical techniques (chromatography, electrophoresis, mass spectrometry, immunology) have been used, either alone or in combination. The main advantages of HPLC analysis lie in its high resolution power and versatility. In a single chromatographic run, it is possible to obtain both the composition and the amount of the protein fraction and analysis can be automated. 

Haq A, Lobo PI, Al-Tufail M, Rama NR, Al-Sedairy ST. Immunomodulatory effect of Nigella sativa proteins fractionated by ion exchange chromatography. International Journal of Immunopharmacology 1999 21 283-295. 

J. Porath, J. Carlsson, I. Olsson, and G. Belfrage, Nature258, 598-599 (1975). Metal Chelate Affinity Chromatography, a New Approach to Protein Fractionation. ... 

AR Hill, B Manji, Y Kakuda, C Myers, DM Irvine. Quantification and characterization of whey protein fractions separated by anion exchange chromatography. Milchwissenschaft 42 693-696, 1987. 

CM Hollar, AJR Law, DG Dagleish, RJ Brown. Separation of major casein fractions using cation-exchange fast protein liquid chromatography. J Dairy Sci 74 2403-2409, 1991. 

P3. Porath, J., Zone Electrophoresis in Columns and Adsorption Chromatography on Ionic Cellulose Derivatives, as Methods for Peptide and Protein Fractionations. Almqvist and Wiksells, Uppsala, 1957. 

The use of these natural fluorescence techniques offers not only the possibility of studying the interaction of proteins with membranes, under convective and diffusive conditions, but also they may be easily extended to studies involving proteins and other porous materials such as chromatography media. The areas of application of these techniques will range from polypeptide and protein fractionation to the monitoring of systems where protein-surface interactions are relevant. 

---