Grunewald, Jan, Jones, David, Brock, Ansgar, Bursulaya, Badry, Patterson, Paula, Ng, Kenneth, Vo, Michael, Uno, Tetsuo, Hunt, James, Spraggon, Glen and Geierstanger, Bernhard (2014) Site-Specific Dual-Labeling of Proteins Using Small Orthogonal Tags at Neutral pH. ChemBiochem.

Abstract

To expand the utility of proteinaceous FRET biosensors, we have developed a dual-labeling approach based on two small bio-orthogonal tags, pyrroline-carboxy-lysine (Pcl) and the S6 peptide. The lack of cross-reactivity between those tags enables site-specific two-color protein conjugation in a one-pot reaction. Moreover, Pcl/S6 dual-tagged proteins can be produced in both bacterial and mammalian expression systems, as demonstrated for Z domain and IgE-Fc, respectively. Both proteins could be efficiently dual-labeled with FRET-compatible fluorescent dyes at neutral pH. In the case of IgE-Fc, the resulting conjugate enabled the monitoring of IgE binding to its high-affinity receptor, FcRI, which is considered a key event in allergic reactions.

Item Type:	Article
Date Deposited:	26 Apr 2016 23:45
Last Modified:	26 Apr 2016 23:45
URI:	https://oak.novartis.com/id/eprint/21617