Alexa L Andrzejewski, Joshua Ferrar, Marla Kratzer-Charron, Mark E Bowen, Ucheor B Choi
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引用次数: 0
Abstract
Single-molecule fluorescence resonance energy transfer (smFRET) is a powerful technique for studying the structural dynamics of protein molecules or detecting interactions between protein molecules in real time. Due to the high sensitivity in spatial and temporal resolution, smFRET can decipher sub-populations within heterogeneous native state conformations, which are generally lost in traditional measurements due to ensemble averaging. In addition, the single-molecule reconstitution allows protein molecules to be observed for an extensive period of time and can recapitulate the geometry of the cellular environment to retain biological function. Here we provide a detailed method of using smFRET to monitor the conformational dynamics of syntaxin-3b from the ribbon synapses during assembly of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex.
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For over 20 years, biological scientists have come to rely on the research protocols and methodologies in the critically acclaimed Methods in Molecular Biology series. The series was the first to introduce the step-by-step protocols approach that has become the standard in all biomedical protocol publishing. Each protocol is provided in readily-reproducible step-by-step fashion, opening with an introductory overview, a list of the materials and reagents needed to complete the experiment, and followed by a detailed procedure that is supported with a helpful notes section offering tips and tricks of the trade as well as troubleshooting advice.
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