Esposito Rosario, Delfino Ines, Portaccio Marianna, Iannuzzi Clara, Lepore Maria
Dipartimento di Ingegneria Chimica, dei Materiali e della Produzione industriale, Università Federico II, P.le Tecchio 80, 80125, Naples, Italy.
Dipartimento di Scienze Ecologiche e Biologiche, Università della Tuscia, Largo dell'Università snc, 01100, Viterbo, Italy.
Eur Biophys J. 2019 May;48(4):395-403. doi: 10.1007/s00249-019-01369-0. Epub 2019 May 4.
Optical properties of flavin adenine dinucleotide (FAD) moiety are widely used nowadays for biotechnological applications. Given the fundamental role played by FAD, additional structural information about this enzymatic cofactor can be extremely useful in order to obtain a greater insight into its functional role in proteins. For this purpose, we have investigated FAD behaviour in aqueous solutions at different pH values by a novel approach based on the combined use of time-resolved fluorescence and circular dichroism spectroscopies. The results showed that pH strongly affects time-resolved fluorescence emission and the analysis allowed us to detect a three-component decay for FAD in aqueous solution with pH-depending lifetimes and relative amplitudes. Circular dichroism data were analyzed by a multi-Gaussian fitting procedure and the trends of properly chosen parameters confirmed pH-depending changes. The comparison between the results obtained by these two optical techniques allowed us to improve the significance of the outcome of circular dichroism. This combined approach may provide a useful tool for biotechnological investigation.
黄素腺嘌呤二核苷酸（FAD）部分的光学性质如今在生物技术应用中得到了广泛应用。鉴于FAD所起的基础作用，关于这种酶辅因子的额外结构信息对于更深入了解其在蛋白质中的功能作用可能极其有用。为此，我们采用了一种基于时间分辨荧光和圆二色光谱联用的新方法，研究了不同pH值下FAD在水溶液中的行为。结果表明，pH值强烈影响时间分辨荧光发射，通过分析我们检测到水溶液中FAD的三组分衰减，其寿命和相对振幅取决于pH值。通过多高斯拟合程序分析圆二色性数据，适当选择的参数趋势证实了随pH值变化的情况。这两种光学技术所得结果的比较使我们能够提高圆二色性结果的显著性。这种联合方法可能为生物技术研究提供一个有用的工具。
