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[86] Difference spectroscopy

Methods in Enzymology, ISSN: 0076-6879, Vol: 11, Issue: C, Page: 748-775
1967
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  • Citations
    185
    • Citation Indexes
      185
  • Captures
    8

Article Description

This chapter focuses on the techniques of difference spectroscopy. Difference spectral methods offer the possibility of dealing experimentally with questions concerning the involvement and location of natural and artificial chromophoric groups in enzyme-substrate and protein-protein interactions. The principle of solvent perturbation technique of difference spectroscopy for studying the location of chromophoric amino acids in proteins is also discussed. The study of lysozyme and α-chymotrypsinogen, the solvent perturbation technique is applied to the study of a number of other enzymes and proteins. It is also used to study proteins and polypeptides in helix-promoting and random coilpromoting organic solvents. Acid- and alkali-induced difference spectra in proteins owe their origin to a number of interrelated physical phenomena. Temperature difference spectra are also described. The main spectral changes observed with tyrosine-containing proteins in the alkaline pH region are due to the ionization of tyrosyl residues. Difference spectral techniques have extensive application in denaturation studies of proteins by means of a variety of denaturing agents and salts.

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