School of Natural Sciences, Macquarie University, Sydney, New South Wales, Australia.
Australian Research Council Industrial Transformation Training Centre for Facilitated Advancement of Australia's Bioactives (FAAB), Sydney, New South Wales, Australia.
Chirality. 2023 Aug;35(8):498-504. doi: 10.1002/chir.23554. Epub 2023 Mar 9.
Membranes are important sites of intermolecular interactions in biological systems. However, they present significant analytical challenges as they contain multiple analytes and are dynamic in nature. In this work, we show how a Jasco J-1500 circular dichroism spectropolarimeter can be used with a microvolume Couette flow cell and appropriate cut-off filters to measure excitation fluorescence detected linear dichroism (FDLD) of fluorophores embedded in liposomal membranes. The result is a spectrum that selectively probes the fluorophore(s) and eliminates the scattering that is apparent in the corresponding flow linear dichroism (LD) spectrum. The FDLD spectrum is opposite in sign from the LD spectrum with relative magnitudes modified by the quantum yields of the transitions. FDLD thus enables analyte orientations to be identified in a membrane. Data for a membrane peptide, gramicidin, and two aromatic analytes, anthracene and pyrene, are presented. Issues with the "leakage" of photons by the long pass filters used is also discussed.
膜是生物系统中分子间相互作用的重要场所。然而，由于它们包含多种分析物且具有动态性质，因此分析它们具有很大的挑战性。在这项工作中，我们展示了如何使用 Jasco J-1500 圆二色光谱仪与微体积 Couette 流动池和适当的截止滤波器来测量嵌入脂质体膜中的荧光团的激发荧光检测线二色性（FDLD）。结果是得到一个选择性探测荧光团并消除在相应流动线二色性（LD）光谱中明显存在的散射的光谱。FDLD 光谱的符号与 LD 光谱相反，相对幅度由跃迁的量子产率修改。因此，FDLD 可以识别膜中的分析物取向。本文提供了一种膜肽（gramicidin）和两种芳族分析物（蒽和芘）的 FDLD 数据。还讨论了长通滤波器中光子“泄漏”的问题。
