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DiB-splits: nature-guided design of a novel fluorescent labeling split system
Nina G Bozhanova1, Alexey S Gavrikov2, Alexander S Mishin2
1Department of Chemistry, Center for Structural Biology, Vanderbilt University, Nashville, TN, 37235, USA.
Scientific Reports
|July 8, 2020
Summary
Researchers developed DiB-splits, a smaller, self-assembling fluorogen-activating protein (FAP) system. This innovative probe offers tunable fluorescence and maintains key properties for advanced microscopy and sensor development.
Area of Science:
- Biochemistry
- Molecular Biology
- Biotechnology
Background:
- Fluorogen-activating proteins (FAPs) combine benefits of genetically encoded proteins and external fluorogens for tunable fluorescence.
- Previous FAPs, like green- and red-emitting DiBs derived from bacterial lipocalin Blc, offered on-demand signaling.
Purpose of the Study:
- To design and characterize the first self-assembling FAP split system, termed DiB-splits.
- To reduce the size of FAP labels while retaining desirable fluorescence properties.
Main Methods:
- Rational design of a split FAP system.
- Functional characterization of the DiB-splits system.
- Structural characterization of the DiB-splits system.
Main Results:
- Developed DiB-splits, a self-assembling FAP split system with reduced label size (~8-12 kDa).
- DiB-splits preserve high fluorescence intensity increase upon chromophore binding.
- Achieved nanomolar to low micromolar binding affinities and high photostability.
Conclusions:
- DiB-splits offer a smaller, versatile FAP label suitable for various microscopy techniques (wide-field, confocal, super-resolution).
- This system serves as a foundation for developing protein-protein interaction detection and novel FAP-based sensors.
