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1). Fig. 1. Isoelectric focusing of a mixture of proteins with pI values 4, 5, 6 and 7. After the pH gradient is established by the electric field, the proteins migrate and focus at their pI values. B3 – Protein analysis 27 Hydrophobicity Hydrophobic interactions between proteins and a column material containing aromatic or aliphatic alkyl groups are promoted in solutions of high ionic strength. By applying a gradient of decreasing ionic strength, proteins elute at different stages. Affinity The high specificities of enzyme–substrate, receptor–ligand and antibody–antigen interactions mean that these can be exploited to give very high degrees of purification.

Tertiary structure The different sections of secondary structure and connecting regions fold into a well-defined tertiary structure, with hydrophilic amino acids mostly on the surface and hydrophobic ones in the interior. The structure is stabilized by noncovalent interactions and sometimes disulfide bonds. Denaturation leads to loss of secondary and tertiary structure. Quaternary structure Many proteins have more than one polypeptide subunit. Hemoglobin has two ␣ and two ␤ chains. Large complexes such as microtubules are constructed from the quaternary association of individual polypeptide chains.

RNA is also susceptible to hydrolysis at high pH, by participation of the 2Ј-OH in intramolecular cleavage of the phosphodiester backbone. Some chemicals, such as urea and formamide, can denature DNA and RNA at neutral pH by disrupting the hydrophobic forces between the stacked bases. DNA is very long and thin, and DNA solutions have a high viscosity. Long DNA molecules are susceptible to cleavage by shearing in solution – this process can be used to generate DNA of a specific average length. 7 g cm–3, and can be analyzed and purified by its ability to equilibrate at its buoyant density in a cesium chloride density gradient formed in a centrifuge.