Download HPLC of Peptides and Proteins: Methods and Protocols by Marie-Isabel Aguilar PDF

By Marie-Isabel Aguilar

Monash Univ., Clayton, Victoria, Australia. presents entire insurance of all popular HPLC ideas with step by step directions. provides examples of a variety of peptide and protein functions and discusses key separation parameters for reaching excessive answer. For researchers. DNLM: Peptides--chemistry.

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Additional info for HPLC of Peptides and Proteins: Methods and Protocols (Methods in Molecular Biology)

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As mentioned earlier (see Subheading 1. and Fig. 1), the charge on a protein is pH dependent. Hence, once bound to the IEX support, alteration of the pH of the eluent can be used to change the charge on the protein and bring it closer to its pI, causing elution of the protein from the ion-exchanger. However, it is difficult to produce a linear pH gradient at a constant salt concentration, and for this reason, pH gradients are not as common as salt gradients as a means to elute proteins in ion-exchange chromatography (4).

Chromatogr. 499, 89–102. 7. Unger, K. K. (1990) Silica as a support, in HPLC of Biological Macromolecules: Methods and Applications (Gooding, K. M. and Regnier, F. ), Dekker, New York, pp. 3–24. 8. Henry, M. (1991) Design requirements of silica-based matrices for biopolymer chromatography. J. Chromatogr. 544, 413–443. RP-HPLC 21 9. Zhou, N. , Mant, C. , Kirkland J. , and Hodges R. S. (1991) Comparison of silica-based cyanopropyl and octyl reversed phase packings for the separation of peptides and proteins.

The disadvantages of IEX are few, but include 1) the sample must be applied to the IEX support under conditions of low ionic strength and controlled pH, which sometimes requires an extra buffer exchange step to be inserted, 2) chromatographic instrumentation should be resistant to salt-induced corrosion, and 3) postchromatographic concentration of dilute solutions of recovered proteins can result in high salt concentrations (>1 M), unsuitable, for example, in biological assays unless buffer exchange is carried out.

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