title: Crystal structure of the cyan fluorescent protein Cerulean-S175G.
abstract: Enhanced cyan fluorescent protein (ECFP) was derived from Aequorea victoria green fluorescent protein (avGFP), notably with S65T/Y66W mutations. Its chromophore consists of a tripeptide comprised of Thr65, Trp66 and Gly67 (TWG) residues, while that of avGFP consists of a Ser65, Tyr66 and Gly67 (SYG) tripeptide. Cerulean and SCFP3A were derived from ECFP-S72A/H148D (a double mutation) with additional Y145A and S175G mutations, respectively, while Cerulean-S175G has both mutations (Y145A and S175G). The crystal structures of these ECFP variants at neutral pH were reported to adopt two distinct major conformations called ECFP and Cerulean. In this study, Cerulean-S175G was revealed to adopt only the Cerulean conformation, while Cerulean has been reported to adopt both the ECFP and the Cerulean conformations in its crystal structures. Sharing the same S175G mutation with SCFP3A, Cerulean-S175G showed a slightly increased quantum yield, like SCFP3A, but did not adopt the ECFP conformation adopted by SCFP3A. Detailed comparison of Cerulean-S175G and other ECFP variants revealed that the notable conformational changes in ECFP variants can be understood mainly in terms of the interaction between the Trp66 residue of the chromophore and residues 145-148 of β-strand 7.
authors.affiliation: Bio-Analytical Science Major, Korea University of Science and Technology, KRIBB Campus, 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.
authors.forename: Sang Wook
authors.initials: SW
authors.lastname: Park
authors.affiliation: Bio-Analytical Science Major, Korea University of Science and Technology, KRIBB Campus, 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.
authors.forename: Sunghyun
authors.initials: S
authors.lastname: Kang
authors.affiliation: Bio-Analytical Science Major, Korea University of Science and Technology, KRIBB Campus, 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.
authors.forename: Tae Sung
authors.initials: TS
authors.lastname: Yoon
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D015870:Gene Expression; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D037402:Hydrozoa / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D020033:Protein Isoforms; D011994:Recombinant Proteins; C527701:enhanced cyan fluorescent protein; D049452:Green Fluorescent Proteins
keywords: Cerulean-S175G; enhanced cyan fluorescent protein; fluorescence lifetime imaging microscopy; fluorescence resonance energy transfer; fluorophores