title: Orange fluorescent proteins constructed from cyanobacteriochromes chromophorylated with phycoerythrobilin.
abstract: Cyanobacteriochromes are a structurally and spectrally highly diverse class of phytochrome-related photosensory biliproteins. They contain one or more GAF domains that bind phycocyanobilin (PCB) autocatalytically; some of these proteins are also capable of further modifying PCB to phycoviolobilin or rubins. We tested the chromophorylation with the non-photochromic phycoerythrobilin (PEB) of 16 cyanobacteriochrome GAFs from Nostoc sp. PCC 7120, of Slr1393 from Synechocystis sp. PCC 6803, and of Tlr0911 from Thermosynechococcus elongatus BP-1. Nine GAFs could be autocatalytically chromophorylated in vivo/in E. coli with PEB, resulting in highly fluorescent biliproteins with brightness comparable to that of fluorescent proteins like GFP. In several GAFs, PEB was concomitantly converted to phycourobilin (PUB) during binding. This not only shifted the spectra, but also increased the Stokes shift. The chromophorylated GAFs could be oligomerized further by attaching a GCN4 leucine zipper domain, thereby enhancing the absorbance and fluorescence of the complexes. The presence of both PEB and PUB makes these oligomeric GAF-"bundles" interesting models for energy transfer akin to the antenna complexes found in cyanobacterial phycobilisomes. The thermal and photochemical stability and their strong brightness make these constructs promising orange fluorescent biomarkers.
authors.affiliation: State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, P.R. China. zhouming321@126.com.
authors.forename: Ya-Fang
authors.initials: YF
authors.lastname: Sun
authors.forename: Jin-Guo
authors.initials: JG
authors.lastname: Xu
authors.forename: Kun
authors.initials: K
authors.lastname: Tang
authors.forename: Dan
authors.initials: D
authors.lastname: Miao
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Gärtner
authors.forename: Hugo
authors.initials: H
authors.lastname: Scheer
authors.forename: Kai-Hong
authors.initials: KH
authors.lastname: Zhao
authors.forename: Ming
authors.initials: M
authors.lastname: Zhou
mesh_terms: D000458:Cyanobacteria / Q000737:chemistry / Q000378:metabolism*; D008164:Luminescent Proteins / Q000737:chemistry / Q000378:metabolism*; D055668:Photochemical Processes; D052980:Phycobilins / Q000737:chemistry / Q000378:metabolism*; D010799:Phycoerythrin / Q000737:chemistry / Q000378:metabolism*; D013696:Temperature; D014557:Urobilin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
chemical_list: D008164:Luminescent Proteins; D052980:Phycobilins; D010799:Phycoerythrin; C011070:phycoerythrobilin; D014557:Urobilin; C044986:phycourobilin