The Quadricyclane Ligation, a reaction between quadricyclane (QC) and nickel-bis(dithiolene), possesses many of the requisite qualities of a bioorthogonal labeling technique. The reaction proceeds rapidly in aqueous environments at physiological pH and temperature, and occurs in the presence of complex cell lysate. In this work, we aim to expand the utility of the QC Ligation as a bioorthogonal conjugation strategy by pursuing 1) site-specific incorporation of the QC functionality into a protein of interest and 2) optimization of the ligation chemistry to enhance signal over background in in vitro labeling experiments. To address the first goal, we have synthesized QC-containing amino acid 1  and have incorporated it into a superfolder GFP using the pyrrolysine synthetase (PylRS) tRNACUA system. This work represents the first example of the site-specific incorporation of QC into a protein. We next show that the QC-containing GFP can be selectively labeled with Ni-bis(dithiolene) 2 in the presence of complex cell lysate. Currently, we are working to optimize the chemistry of the QC-ligation by exploring the reactivities of various metal�bis(dithiolene) analogs. Specifically, we are investigating the utility of palladium-bis(dithiolenes), which we have found to react with QC an order of magnitude faster than the corresponding Ni compound.

Development of the quadricyclane ligation as a bioorthogonal labeling strategy

248th National Meeting (2014)

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