title: A generic building block for C- and N-terminal protein-labeling and protein-immobilization.
abstract: Expressed protein ligation (EPL) and bioconjugation based on the maleimide group (MIC-conjugation) provide powerful tools for protein modification. In the light of the importance of site-selectively modified proteins for the study of protein function, a flexible method for the introduction of tags and reporter groups into the C-terminus of proteins employing EPL and MIC-conjugation was developed. We describe the solid-phase synthesis of a generic building block, equipped with fluorescence markers or different functional groups. This generic building block allows for a flexible incorporation of different tags into proteins and was used for the introduction of fluorescence markers into the C-terminus of Rab and Ras GTPases by EPL or MIC-conjugation techniques. In addition, a building block appropriately modified for the incorporation of an azide into proteins was synthesized. Azide-functionalized Ras protein was immobilized on a phosphane-modified surface by means of Staudinger ligation providing a highly chemoselective ligation method for the immobilization of proteins.
authors.affiliation: Max-Planck-Institut für molekulare Physiologie, Abteilung Chemische Biologie, Otto-Hahn-Strasse 11, 44227 Dortmund, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Watzke
authors.forename: Marta
authors.initials: M
authors.lastname: Gutierrez-Rodriguez
authors.forename: Maja
authors.initials: M
authors.lastname: Köhn
authors.forename: Ron
authors.initials: R
authors.lastname: Wacker
authors.forename: Hendrik
authors.initials: H
authors.lastname: Schroeder
authors.forename: Rolf
authors.initials: R
authors.lastname: Breinbauer
authors.forename: Jürgen
authors.initials: J
authors.lastname: Kuhlmann
authors.forename: Kirill
authors.initials: K
authors.lastname: Alexandrov
authors.forename: Christof M
authors.initials: CM
authors.lastname: Niemeyer
authors.forename: Roger S
authors.initials: RS
authors.lastname: Goody
authors.forename: Herbert
authors.initials: H
authors.lastname: Waldmann
mesh_terms: D001386:Azides / Q000737:chemistry; D001665:Binding Sites; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D019204:GTP-Binding Proteins / Q000737:chemistry*; D008301:Maleimides / Q000138:chemical synthesis / Q000737:chemistry*; D015394:Molecular Structure; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D011506:Proteins / Q000737:chemistry*; D013329:Structure-Activity Relationship; D018631:ras Proteins / Q000737:chemistry
chemical_list: D001386:Azides; D005456:Fluorescent Dyes; D008301:Maleimides; D011506:Proteins; D019204:GTP-Binding Proteins; D018631:ras Proteins