title: Generation of a genetically encoded, photoactivatable intein for the controlled production of cyclic peptides.
abstract: Cyclic peptides are important natural products and hold great promise for the identification of new bioactive molecules. The split-intein-mediated SICLOPPS technology provides a generic access to fully genetically encoded head-to-tail cyclized peptides and large libraries thereof (SICLOPPS=split-intein circular ligation of peptides and proteins). However, owing to the spontaneous protein splicing reaction, product formation occurs inside cells, making peptide isolation inconvenient and precluding traditional in vitro assays for inhibitor discovery. The design of a genetically encoded, light-dependent intein using the photocaged tyrosine derivative ortho-nitrobenzyltyrosine incorporated at an internal, non-catalytic position is now reported. Stable intein precursors were purified from the E.coli expression host and subsequently subjected to light activation in vitro for both the regular protein splicing format and cyclic peptide production, including the natural product segetalin H as an example. The activity of the intein could also be triggered in living cells.
authors.affiliation: Department Chemistry and Pharmacy, Institute of Biochemistry, University of Muenster, Wilhelm-Klemm-Strasse 2, 48149 Münster (Germany).
authors.forename: Jana K
authors.initials: JK
authors.lastname: Böcker
authors.forename: Kristina
authors.initials: K
authors.lastname: Friedel
authors.forename: Julian C J
authors.initials: JC
authors.lastname: Matern
authors.forename: Anne-Lena
authors.initials: AL
authors.lastname: Bachmann
authors.forename: Henning D
authors.initials: HD
authors.lastname: Mootz
mesh_terms: D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D047668:Inteins; D008027:Light; D008969:Molecular Sequence Data; D019151:Peptide Library; D010456:Peptides, Cyclic / Q000737:chemistry / Q000235:genetics*; D055668:Photochemical Processes; D015202:Protein Engineering; D019154:Protein Splicing; D014443:Tyrosine / Q000031:analogs & derivatives / Q000235:genetics
chemical_list: D019151:Peptide Library; D010456:Peptides, Cyclic; C105633:segetalin H; D014443:Tyrosine
keywords: macrocycles; peptides; photocontrol; protein engineering; protein splicing