title: Multicolor Imaging of Bifacial Activities of Estrogens.
abstract: The present protocol introduces multicolor imaging of bifacial activities of an estrogen. For the multicolor imaging, the authors fabricated two single-chain probes emitting green or red bioluminescence (named Simer-G and -R, respectively) from click beetle luciferase (CBLuc) green and red: Simer-R consists of the ligand binding domain of estrogen receptor (ER LBD) and the Src homology-2 (SH2) domain of Src, which are sandwiched between split-CBLuc red (CBLuc-R). On the other hand, Simer-G emitting red light consists of the ER LBD and a common consensus sequence of coactivators (LXXLL motif), which are inserted between split-CBLuc green (CBLuc-G). This probe set creates fingerprinting spectra from the characteristic green and red bioluminescence in response to agonistic and antagonistic activities of a ligand of interest. The present protocol further provides a unique methodology to calculate characteristic estrogenicity scores of various ligands from the spectra.
authors.affiliation: Research Institute for Environmental Management Technology, National Institute of Advanced Industrial Science and Technology (AIST), 16-1 Onogawa, Tsukuba, 305-8569, Japan. kimu-sb@aist.go.jp.
authors.forename: Sung-Bae
authors.initials: SB
authors.lastname: Kim
authors.affiliation: Department of Chemistry, School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Umezawa
mesh_terms: D000818:Animals; D002460:Cell Line; D004967:Estrogens / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D023061:Gene Order; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D008024:Ligands; D008163:Luminescent Measurements / Q000379:methods; D057054:Molecular Imaging / Q000379:methods*; D015335:Molecular Probes / Q000737:chemistry / Q000235:genetics; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D011960:Receptors, Estrogen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004967:Estrogens; D008024:Ligands; D015335:Molecular Probes; D011960:Receptors, Estrogen
keywords: Bioluminescence; Click beetle; Estrogen receptor; Luciferase; Multicolor imaging; Single-chain probe