Utility of 5-Cyanotryptophan Fluorescence as a Sensitive Probe of Protein Hydration

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• 作者：Beatrice N. Markiewicz ; Debopreeti Mukherjee ; Thomas Troxler ; Feng Gai
• 刊名：Journal of Physical Chemistry B
• 出版年：2016
• 出版时间：February 11, 2016
• 年：2016
• 卷：120
• 期：5
• 页码：936-944
• 全文大小：460K
• ISSN：1520-5207

文摘

Tryptophan (Trp) fluorescence has been widely used to interrogate the structure, dynamics, and function of proteins. In particular, it provides a convenient and site-specific means to probe a protein’s hydration status and dynamics. Herein, we show that a tryptophan analogue, 5-cyanotryptophan (Trp_CN), can also be used for this purpose, but with the benefit of enhanced sensitivity to hydration. This conclusion is reached based on measurements of the static and time-resolved fluorescence properties of 5-cyanoindole, Trp_CN, and Trp_CN-containing peptides in different solvents, which indicate that upon dehydration the fluorescence quantum yield (QY) and lifetime (τ_F) of Trp_CN undergo a much greater change in comparison to those of Trp. For example, in H₂O the QY of Trp_CN is less than 0.01, which increases to 0.11 in 1,4-dioxane. Consistently, the fluorescence decay kinetics of Trp_CN in H₂O are dominated by a 0.4 ns component, whereas in 1,4-dioxane the kinetics are dominated by a 6.0 ns component. The versatile utility of Trp_CN as a sensitive fluorescence reporter is further demonstrated in three applications, where we used it (1) to probe the solvent property of a binary mixture consisting of dimethyl sulfoxide and H₂O, (2) to monitor the binding interaction of an antimicrobial peptide with lipid membranes, and (3) to differentiate two differently hydrated environments in a folded protein.