A method to probe protein structure from UV absorbance spectra

Anal Biochem. 2019 Dec 15;587:113450. doi: 10.1016/j.ab.2019.113450. Epub 2019 Sep 21.


Proteins primarily absorb UV light due to the presence of tryptophan, tyrosine, and phenylalanine residues, with absorbance maxima at 280, 275, and 258 nm, respectively. We now demonstrate that a simple value obtained by relating the absorbance at all three wavelengths, [A280/A275 + A280/A258], is a generally useful, robust, and sensitive probe of protein 'foldedness', and thus can be used to investigate unfolding, refolding, disulfide bonds, stability, buffer excipients, and even protein-protein and protein-ligand interactions.

Keywords: Protein conformation; Protein denaturation; Protein folding; Protein misfolding; Protein stability; Protein structure; Protein-protein interaction; UV–Vis spectroscopy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aspartic Acid Proteases / chemistry*
  • Aspartic Acid Proteases / metabolism
  • Hydrogen-Ion Concentration
  • Pepsin A / chemistry*
  • Pepsin A / metabolism
  • Protein Conformation
  • Protein Folding
  • Spectrophotometry, Ultraviolet
  • Ultraviolet Rays*


  • Aspartic Acid Proteases
  • Pepsin A