title: Click-tag and amine-tag: chemical tag approaches for efficient protein labeling in vitro and on live cells using the naturally split Npu DnaE intein.
abstract: Protein labeling with synthetic moieties remains in many cases a technically challenging or unresolved task. Two new and simple concepts are presented. In both approaches, a very short tag of only a few amino acids is prepared with the desired chemical modification and, in a second step, it is transferred to the protein of interest by protein trans-splicing. For the amine-tag, a recombinant intein fragment free of lysine residues was generated such that the amine group of the N terminus could be selectively modified with regular amine-reactive reagents. Thus, standard bioconjugation procedures without any chemical synthesis could be applied without modification of lysines in the protein of interest. For the click-tag, protein trans-splicing was combined with unnatural amino acid mutagenesis and subsequent bioorthogonal side chain modification, as demonstrated for click chemistry using p-azidophenylalanine. By the two-step strategy, exposure of the protein of interest to the copper catalyst was avoided.
authors.affiliation: Institute of Biochemistry, University of Muenster, Wilhelm-Klemm-Strasse 2, 48149 Münster (Germany).
authors.forename: Vivien
authors.initials: V
authors.lastname: Schütz
authors.forename: Henning D
authors.initials: HD
authors.lastname: Mootz
mesh_terms: D000588:Amines / Q000737:chemistry*; D006801:Humans; D066298:In Vitro Techniques / Q000379:methods*; D047668:Inteins / Q000235:genetics; D019154:Protein Splicing / Q000235:genetics*; D011506:Proteins / Q000737:chemistry*
chemical_list: D000588:Amines; D011506:Proteins
keywords: bioconjugation; bioorthogonality; fusion tags; lysine modification; unnatural amino acids