title: Semi-synthesis and analysis of chemically modified zif268 zinc-finger domains.
abstract: Total synthesis of proteins can be challenging despite assembling techniques, such as native chemical ligation (NCL) and expressed protein ligation (EPL). Especially, the combination of recombinant protein expression and chemically addressable solid-phase peptide synthesis (SPPS) is well suited for the redesign of native protein structures. Incorporation of analytical probes and artificial amino acids into full-length natural protein domains, such as the sequence-specific DNA binding zinc-finger motifs, are of interest combining selective DNA recognition and artificial function. The semi-synthesis of the natural 90 amino acid long sequence of the zinc-finger domain of Zif268 is described including various chemically modified constructs. Our approach offers the possibility to exchange any amino acid within the third zinc finger. The realized modifications of the natural sequence include point mutations, attachment of a fluorophore, and the exchange of amino acids at different positions in the zinc finger by artificial amino acids to create additional metal binding sites. The individual constructs were analyzed by circular dichroism (CD) spectroscopy with respect to the integrity of the zinc-finger fold and DNA binding.
authors.affiliation: Georg-August-Universität Göttingen, Institut für Organische und Biomolekulare Chemie Tammannstrasse 2, 37077 Göttingen (Germany), E-mail: udieder@gwdg.de.
authors.forename: Friederike
authors.initials: F
authors.lastname: Fehr
authors.affiliation: Georg-August-Universität Göttingen, Institut für Organische und Biomolekulare Chemie Tammannstrasse 2, 37077 Göttingen (Germany), E-mail: udieder@gwdg.de.
authors.forename: André
authors.initials: A
authors.lastname: Nadler
authors.affiliation: Georg-August-Universität Göttingen, Albrecht-von-Haller Institut für Pflanzenwissenschaften Justus-von-Liebig-Weg 11, 37077 Göttingen (Germany).
authors.forename: Florian
authors.initials: F
authors.lastname: Brodhun
authors.affiliation: Georg-August-Universität Göttingen, Albrecht-von-Haller Institut für Pflanzenwissenschaften Justus-von-Liebig-Weg 11, 37077 Göttingen (Germany).
authors.forename: Ivo
authors.initials: I
authors.lastname: Feussner
authors.affiliation: Georg-August-Universität Göttingen, Institut für Organische und Biomolekulare Chemie Tammannstrasse 2, 37077 Göttingen (Germany), E-mail: udieder@gwdg.de.
authors.forename: Ulf
authors.initials: U
authors.lastname: Diederichsen
keywords: expressed protein ligation; metalloproteins; peptide synthesis; proteins; zinc-finger domains