How do reducing agents denature proteins

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The denaturation of proteins

Denaturation is understood to be all physical or chemical processes that change a native protein in such a way that it loses its biological activity or its specific properties, although the primary structure remains unchanged.

Denaturation dissolves hydrogen bonds, hydrophobic interactions and, in the presence of reducing agents, also disulfide bridges. In most cases, the result is an aggregate formation of the proteins up to the precipitation of the protein from the solution, since in the denatured proteins the hydrophobic amino acids normally located inside come into contact with the surrounding (aqueous) solvent and with the hydrophobic amino acids of other proteins interact.

When denaturation is due to the breaking of non-covalent bonds, it is usually reversible. The interaction with the denaturant stabilizes a new conformation, which can also return to the original, native conformation of the protein if the denaturant is removed (renaturation). In principle, disulfide bridges can also be restored.

Irreversible, i.e. irreversible, denaturation occurs when unspecific oxidations of the protein occur or other covalent modifications occur. In this state, the protein takes on a disordered coil structure (random coil) and can no longer be renatured.

The denaturation of a protein leads to versatile physical and chemical changes. Among other things, there is a reduction in the hydration of the protein, solubility and crystallizability change and the isoelectric point shifts (because additional ionizable groups occur). In addition, there is increased UV absorption as more phenolic groups are exposed.

Physical and chemical denaturation of proteins can be achieved, among other things, by:

  • physical denaturation
  • chemical denaturation
  • vigorous stirring, shaking or heating
  • UV, X-ray or radiation absorption
  • Ultrasound treatment
  • 6-8 M urea solution (dissolves hydrogen bonds)
  • 4 M solution of guanidine
  • extreme pH values ​​(below 3 or above 9)
  • Detergents