1. Fluorescent Labeling of Outer Membrane Proteins Using the SpyCatcher-SpyTag System.
- Author
-
Duodu R, Linke D, and Leo JC
- Subjects
- Coloring Agents, Membrane Proteins genetics, Peptides chemistry
- Abstract
The SpyCatcher-SpyTag system has become a popular and versatile tool for protein ligation. It is based on a small globular protein (SpyCatcher) that binds to a 13-residue peptide (SpyTag), which subsequently leads to the formation of a covalent isopeptide bond. Thus, the reaction is essentially irreversible. Here, we describe how the SpyCatcher-SpyTag system can be used to label surface-exposed bacterial outer membrane proteins, e.g., for topology mapping or fluorescent time-course experiments. We cover using fluorescence measurements and microscopy to measure labeling efficiency using SpyCatcher fused with superfolder GFP in this chapter., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
- Published
- 2024
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