Open AccessRole of conformational changes in the elution of proteins from reversed-phase HPLC columns.
Author(s) -
Gil E. Katzenstein,
Susan Vrona,
R J Wechsler,
Bryan L. Steadman,
Randolph V. Lewis,
C.Russel Middaugh
Publication year - 1986
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.83.12.4268
Subject(s) - elution , chemistry , chromatography , fluorescence , high performance liquid chromatography , alkyl , phase (matter) , deconvolution , reversed phase chromatography , conformational isomerism , analytical chemistry (journal) , molecule , organic chemistry , physics , quantum mechanics , algorithm , computer science
To test the hypothesis that conformational alterations might be involved in the elution of proteins from reversed-phase HPLC columns, the conformations of proteins bound onto a C-8 alkyl-bonded silica surface have been examined in the presence of increasing concentrations of the commonly employed eluent, 1-propanol. Using a combination of photoacoustic, diffuse reflectance deconvolution Fourier transform infrared and front face fluorescence spectroscopic techniques (to minimize interference from light scattering), the existence of surface-associated protein conformational changes induced by propanol is unequivocally demonstrated. The linear relationship found between the amount of propanol needed to elute proteins from C-8 columns and the midpoint of spectrally observed structural transitions is consistent with a role for conformational changes in the elution process.