Probing structural features of water-insoluble proteins by front-face fluorescence.
Front-face fluorescence was used to assess some structural features of proteins in wheat flours, taking advantage of the fact that this technique allowed a direct approach to a number of systems in which most proteins cannot be solubilized without using conditions that strongly modify the structure of proteins in the original material. In this study, we addressed structural changes ensuing from solvation and from mechanical deformation of proteins in wheat flour. Solvation of proteins in the systems under investigation was monitored by taking fluorescence emission spectra of the protein tryptophans (or of protein-bound 1,8-anilino-naphthalene-sulfonate added in appropriate amounts) in mixtures of increasing water content. In separate experiments, changes in overall protein surface hydrophobicity were estimated by titrating flours-mixed with appropriate amounts of water to a dough-like consistency-with increasing concentrations of the fluorescent hydrophobic probe, 1,8-anilino-naphthalene-sulfonate. This approach allowed concomitant determination of the overall binding capacity and of the apparent affinity for the probe of proteins. To test the usefulness of the structural information obtained from these approaches, they were also applied to characterize dough at different levels of mechanical stress, prepared from either wheat flour or semolina.
Bonomi
Francesco
F
DISMA, Sezione di Biochimica, University of Milan, Via Celoria 2, Milan I-20133, Italy. francesco.bonomi@unimi.it
Mora
Giuseppe
G
Pagani
Maria Ambrogina
MA
Iametti
Stefania
S
D000817:Anilino Naphthalenesulfonates / Q000737:chemistry; D002523:Edible Grain / Q000737:chemistry; D005453:Fluorescence; D057927:Hydrophobic and Hydrophilic Interactions; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D012016:Reference Values; D012995:Solubility; D017186:Titrimetry; D014357:Trypsin; D014867:Water / Q000737:chemistry*
D000817:Anilino Naphthalenesulfonates; D011506:Proteins; D014867:Water; C027132:1-anilino-8-naphthalenesulfonate; D014357:Trypsin