Related Experiment Videos
Early conformational changes and activity modulation induced by guanidinium chloride on intestinal alkaline
G A Miggiano1, A Mordente, M G Pischiutta
1Istituto di Chimica Biologica, Università Cattolica del Sacro Cuore, Facoltà di Medicina e Chirurgia Agostino Gemelli, Roma, Italy.
Abstract:
Moderate concentrations of guanidinium chloride induce both instantaneous and time-dependent modifications of the catalytic and optical properties of intestinal alkaline phosphatase, which undergoes consecutive conformational transitions at about 0.05 M, 0.25 M and 1.0 M denaturant. A paradoxical activation is observed up to 1.0 M-guanidine, with a maximum at 0.25 M- and a mid-point around 0.5 M-guanidine. Difference absorbance and fluorescence spectra imply a change in the state of ionization of the protein residues, with variation in molecular size suggested by light-scattering. Random-coil formation is indicated by a lower fluorescence yield, a more polar environment of the aromatic residues and another separate tryptophan emission. Iodide quenching confirms the alterations of conformation. Deprotonation favours the loss of the intramolecular constraints and the enhancement of the structure disruption by guanidine.