# Effects of sucrose on the internal dynamics of azurin

 Title Effects of sucrose on the internal dynamics of azurin Publication Type Journal Article Year of Publication 2005 Authors Cioni, P, Bramanti, E, Strambini, GB Journal Biophysical Journal Volume 88 Pagination 4213–4222 ISSN 00063495 (ISSN) Keywords Acrylamide, Article, Azurin, Biophysics, Buffers, concentration response, Drug Stability, Fluorescence, Mutagenesis, mutant protein, nonhuman, Protein Denaturation, Protein Folding, Protein Structure, Pseudomonas aeruginosa, Recombinant Proteins, Site-Directed, Spectrometry, Sucrose, Temperature, thermodynamics, Trehalose, Xylitol Abstract Sucrose is a natural osmolyte accumulated in cells of organisms as they adapt to environmental stresses. In vitro, sucrose increases protein stability and forces partially unfolded structures to refold. Its effects on the native fold structure and dynamics are not fully established. This study, utilizing Trp phosphorescence spectroscopy, examined the influence of molar concentrations of sucrose on the flexibility of metal-free azurin from Pseudomonas aeruginosa. In addition, by means of specific mutants of the test protein, namely I7S, F110S, and C3A/C26A, that altered its thermodynamic stability, its intrinsic flexibility, and the extent of internal hydration, this investigation sought to identify possible correlations between these features of protein structure and the influence of the osmolyte on protein dynamics. Alterations of structural fluctuations were assessed by both the intrinsic phosphorescence lifetime ($\tau$), which reports on local structure about the triplet probe, and the acrylamide bimolecular quenching rate constant (kq) that is a measure of the average acrylamide diffusion coefficient through the macromolecule. From the modulation of $\tau$ and kq across a wide temperature range and up to a concentration of 2M sucrose, it is concluded that sucrose attenuates structural fluctuations principally when macromolecules are internally hydrated and thermally expanded. Preliminary tests with trehalose and xylitol suggest that the effects of sucrose are general of the polyol class of osmolytes. © 2005 by the Biophysical Society. URL http://www.scopus.com/inward/record.url?eid=2-s2.0-22244455728&partnerID=40&md5=8d1ed52c4b31354033ef8eafacad28ef DOI 10.1529/biophysj.105.060517