A Bimane‐Based Peptide Staple for Combined Helical Induction and Fluorescent Imaging

The thiol‐selective fluorescent imaging agent, dibromobimane, has been repurposed to crosslink cysteine‐ and homocysteine‐containing peptides, with the resulting bimane linker acting as both a structural constraint and a fluorescent tag. Macrocyclisation was conducted on nine short peptides containi...

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Bibliographic Details
Published in:Chembiochem : a European journal of chemical biology 2020-12, Vol.21 (23), p.3423-3432
Main Authors: Horsfall, Aimee J., Dunning, Kylie R., Keeling, Kelly L., Scanlon, Denis B., Wegener, Kate L., Abell, Andrew D.
Format: Article
Language:English
Subjects:
Aqueous solutions
Atomic structure
bimane
Constraints
Crosslinking
Cytosol
Fluorescence
Fluoroscopic imaging
helical structures
Homocysteine
NMR
Nuclear magnetic resonance
peptide staple
Peptides
peptidomimetics
Protein structure
Secondary structure
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Summary:The thiol‐selective fluorescent imaging agent, dibromobimane, has been repurposed to crosslink cysteine‐ and homocysteine‐containing peptides, with the resulting bimane linker acting as both a structural constraint and a fluorescent tag. Macrocyclisation was conducted on nine short peptides containing two cysteines and/or homocysteines, both on‐resin and in buffered aqueous solution, to give macrocycles ranging in size from 16 (i,i+2) to 31 (i,i+7) atoms. The structures were defined by CD, NMR structure calculations by using Xplor‐NIH, NMR secondary shift and JHαNH analyses to reveal helical structure in the i,i+4 (1, 2), and i,i+3 (5) constrained peptides. Cellular‐uptake studies were conducted with three of the macrocycles. Subsequent confocal imaging revealed punctate fluorescence within the cytosol indicative of peptides trapped in endocytic vesicles. These studies demonstrate that dibromobimane is an effective tool for defining secondary structure within short peptides, whilst simultaneously introducing a fluorescent tag suitable for common cell‐based experiments. Two in one: A repurposed fluorescent thiol labelling agent, dibromobimane, is used to form a peptide constraint to introduce helical structure into short peptides. The cyclisation can be conducted on‐resin following SPPS or in solution. The resultant inherently fluorescent peptides are compatible with cell‐uptake imaging experiments and mitigate the need to introduce a secondary tag.
ISSN:1439-4227
1439-7633
DOI:10.1002/cbic.202000485