Effects of pH and salt environment on the association of beta-lactoglobulin revealed by intrinsic fluorescence studies.
The effects of pH, ionic strength and heat on the structure of beta-lactoglobulin (beta-lg) have been investigated by studying the intrinsic tryptophan fluorescence of the protein. Between pH 2 and 9, for sodium chloride concentrations varying from 0.0 to 0.2 M, the position of the fluorescence emission maximum at 20 degrees C remained constant at 328 nm, suggesting that the hydrophobic environment of the fluorophores remained unchanged. The fluorescence intensity increased significantly at pH 2, 7 and 9 on reducing the NaCl concentration of the solutions. The most likely explanation for this, supported by recent light scattering data, is that the presence of NaCl is necessary for beta-lg to dimerize. At the higher NaCl concentrations it was found that dimerization accompanied a reduction in fluorescence intensity. Thus, dissociation appears to reduce the self-quenching of tryptophan residues that occurs within the dimer. The fluorescence of solutions heated below the denaturation temperature reflected the state of association of the protein molecules. Above the denaturation temperature and associated with protein aggregation, an irreversible increase in intrinsic tryptophan fluorescence was observed. We also report what we believe to be the first front-face fluorescence measurements on globular protein gels, showing effects of pH and NaCl concentration.
Renard
D
D
Laboratoire de Physicochimie des Macromolécules, INRA Centre de Recherche de Nantes, France.
Lefebvre
J
J
Griffin
M C
MC
Griffin
W G
WG
D000818:Animals; D002417:Cattle; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007782:Lactoglobulins / Q000187:drug effects / Q000378:metabolism*; D009994:Osmolar Concentration; D011487:Protein Conformation; D012965:Sodium Chloride / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence; D014364:Tryptophan / Q000737:chemistry
D007782:Lactoglobulins; D012965:Sodium Chloride; D014364:Tryptophan