title: Pyrrolysine-inspired protein cyclization.
abstract: The pyrrolysine translational machinery has been extensively explored for the production of recombinant proteins containing a variety of "site-specific" non-canonical amino acids for both in vitro and in vivo biochemical studies. In this study, we report the first use of this technology for the production of branched cyclic proteins with a tadpole-like topology. As a proof of concept, we fused the well-studied RGD peptide to the C terminus of an mCherry reporter protein. Previous studies have shown that cyclization of the RGD peptide enhances its internalization into cells compared to its linear counterpart. The cellular uptake efficiencies of mCherry-cyclo(RGD), mCherry-linear(RGD), and wild-type mCherry determined by flow cytometry follow the trends expected, thereby confirming the feasibility and potential utility of this cyclization approach.
authors.affiliation: The School of Life Sciences, Centre of Novel Biomaterials, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR (China). mariannemmlee@cuhk.edu.hk.
authors.forename: Marianne M
authors.initials: MM
authors.lastname: Lee
authors.forename: Tomasz
authors.initials: T
authors.lastname: Fekner
authors.forename: Jia
authors.initials: J
authors.lastname: Lu
authors.forename: Bradley S
authors.initials: BS
authors.lastname: Heater
authors.forename: Edward J
authors.initials: EJ
authors.lastname: Behrman
authors.forename: Liwen
authors.initials: L
authors.lastname: Zhang
authors.forename: Pang-Hung
authors.initials: PH
authors.lastname: Hsu
authors.forename: Michael K
authors.initials: MK
authors.lastname: Chan
mesh_terms: D003500:Cyclization; D006801:Humans; D008239:Lysine / Q000031:analogs & derivatives* / Q000737:chemistry; D061986:MCF-7 Cells; D009842:Oligopeptides / Q000737:chemistry*; D011487:Protein Conformation; D011506:Proteins / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D009842:Oligopeptides; D011506:Proteins; C047981:arginyl-glycyl-aspartic acid; C456839:pyrrolysine; D008239:Lysine
keywords: RGD; cyclic peptides; integrin; protein engineering; pyrrolysine analogues