Fluorescence and Absorbance Spectroscopy Methods to Study Membrane Perturbations by Antimicrobial Host Defense Peptides
Fluorescence and Absorbance Spectroscopy Methods to Study Membrane Perturbations by Antimicrobial Host Defense Peptides
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DOI：
10.1007/978-1-4939-6737-7_10
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发表时间：
2017-01-01
期刊：
影响因子：
--
通讯作者：
Vogel, Hans J.
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文献类型：
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作者：
Arias, Mauricio;Vogel, Hans J.
抗菌肽（AMP）目前因其作为新型杀菌剂和抑菌剂的潜力而受到广泛研究。许多 AMP 的作用机制涉及细菌膜的透化。已经开发出多种方法来研究肽-膜相互作用；特别是光谱法被广泛使用。含色氨酸 AMP 中色氨酸吲哚环的固有荧光特性可以通过测量荧光蓝移和丙烯酰胺诱导的荧光猝灭来利用。此类研究的一个重要方面是使用不同的细菌膜模型，在大多数情况下是具有不同但明确的磷脂成分的大单层囊泡（LUV）。使用预载有荧光染料（例如钙黄绿素）的 LUV 还可以研究 AMP 的囊泡通透作用。此外，使用基因工程活大肠杆菌细胞进行的实验可用于区分 AMP 对革兰氏阴性菌外膜和内膜的影响。结合起来，这些方法可以提供对 AMP 作用模式的详细了解。
Antimicrobial peptides (AMPs) are currently intensely studied because of their potential as new bactericidal and bacteriostatic agents. The mechanism of action of numerous AMPs involves the permeabilization of bacterial membranes. Several methods have been developed to study peptide-membrane interactions; in particular optical spectroscopy methods are widely used. The intrinsic fluorescence properties of the Trp indole ring in Trp-containing AMPs can be exploited by measuring the fluorescence blue shift and acrylamide-induced fluorescence quenching. One important aspect of such studies is the use of distinct models of the bacterial membrane, in most cases large unilamellar vesicles (LUVs) with different, yet well-defined, phospholipid compositions. Deploying LUVs that are preloaded with fluorescent dyes, such as calcein, also allows for the study of vesicle permeabilization by AMPs. In addition, experiments using genetically engineered live Escherichia coli cells can be used to distinguish between the effects of AMPs on the outer and inner membranes of gram-negative bacteria. In combination, these methods can provide a detailed insight into the mode of action of AMPs.