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“Plug-and-Go” Strategy To Manipulate Streptavidin Valencies

Author(s): Sun, Xun; Montiel, Daniel; Li, Hao; Yang, Haw

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Abstract: The streptavidin-biotin set is one of the most widely utilized conjugation pairs in biotechnological applications. The tetravalent nature of streptavidin and its homologs, however, tends to result in such undesirable complications as crosslinking or ill-defined stoichiometry. Here, we describe a mutagenesis-free strategy to manipulate the valencies of wild-type streptavidin that only requires commercially available reagents. The basic idea is simple: one obtains the desired streptavidin valency by blocking off unwanted binding sites using ancillary biotin (“plug”); this way, the extraordinary fM-biotin-binding affinity is fully retained for the remaining sites in streptavidin. In the present implementation, the ancillary biotin is attached to an auxiliary separation handle, negatively charged DNA or His-tagged protein, via a photochemically or enzymatically cleavable linker. Mixing streptavidin with the ancillary biotin construct produces a distribution of streptavidin valencies. The subsequent chromatographic separation readily isolates the construct of desired streptavidin valency, and the auxiliary handles are easily removed afterwards (“go”). We demonstrate how this “plugand-go” strategy allows a precise control for the compositions of streptavidin-biotin conjugates at the single-molecule level. This low-entry-barrier protocol could further expand the application scope of the streptavidin technology.
Publication Date: 20-Aug-2014
Citation: Sun, Xun, Montiel, Daniel, Li, Hao, Yang, Haw. "“Plug-and-Go” Strategy To Manipulate Streptavidin Valencies" Bioconjugate Chemistry, (8), 25, 1375 - 1380, doi:10.1021/bc500296p
DOI: doi:10.1021/bc500296p
ISSN: 1043-1802
EISSN: 1520-4812
Pages: 1375 - 1380
Type of Material: Journal Article
Journal/Proceeding Title: Bioconjugate Chemistry
Version: This is the author’s final manuscript. All rights reserved to author(s).

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