title: Microbial transglutaminase and c-myc-tag: a strong couple for the functionalization of antibody-like protein scaffolds from discovery platforms.
abstract: Antibody-like proteins selected from discovery platforms are preferentially functionalized by site-specific modification as this approach preserves the binding abilities and allows a side-by-side comparison of multiple conjugates. Here we present an enzymatic bioconjugation platform that targets the c-myc-tag peptide sequence (EQKLISEEDL) as a handle for the site-specific modification of antibody-like proteins. Microbial transglutaminase (MTGase) was exploited to form a stable isopeptide bond between the glutamine on the c-myc-tag and various primary-amine-functionalized substrates. We attached eight different functionalities to a c-myc-tagged antibody fragment and used these bioconjugates for downstream applications such as protein multimerization, immobilization on surfaces, fluorescence microscopy, fluorescence-activated cell sorting, and in vivo nuclear imaging. The results demonstrate the versatility of our conjugation strategy for transforming a c-myc-tagged protein into any desired probe.
authors: Center for Radiopharmaceutical Sciences ETH-PSI-USZ, Paul Scherrer Institute, OIPA10A, 5232 Villigen PSI, (Switzerland).
authors: Patrick
authors: P
authors: Dennler
authors: Laura K
authors: LK
authors: Bailey
authors: Philipp R
authors: PR
authors: Spycher
authors: Roger
authors: R
authors: Schibli
authors: Eliane
authors: E
authors: Fischer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000737:chemistry* / Q000378:metabolism*; D002460:Cell Line; D005260:Female; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D018796:Immunoconjugates / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D016513:Mice, SCID; D015335:Molecular Probes / Q000737:chemistry / Q000378:metabolism; D009374:Neoplasms, Experimental / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D011503:Transglutaminases / Q000378:metabolism*
chemical_list: D000906:Antibodies; D005456:Fluorescent Dyes; D018796:Immunoconjugates; D015335:Molecular Probes; D016271:Proto-Oncogene Proteins c-myc; D011994:Recombinant Proteins; D011503:Transglutaminases
keywords: c-myc-tag; enzymes; microbial transglutaminase; protein modifications; site-specific modification