title: Mutants of Ca2+-regulated Photoprotein Obelin for Site-specific Conjugation.
abstract: Color variants of Ca2+-regulated photoprotein obelin were shown to be an important tool for dual-analyte binding assay. To provide site-directed conjugation with biospecific molecules, several obelin color mutants carrying unique cysteine residues were obtained and characterized for their novel properties. A pair of obelins Y138F,A5C and W92F,H22E,D12C was found to be most suitable (in terms of high bioluminescent activity and stability) as reporters in simultaneous assay of two targets in a sample. Availability of SH-groups, accessible for chemical modification, essentially simplifies the synthesis of biospecific conjugates, increases their yield and conserves obelins' bioluminescence activity. Conjugates with immunoglobulin and oligonucleotide were produced and successfully applied in single nucleotide polymorphism genotyping.
authors.affiliation: Institute of Biophysics SB RAS, Federal Research Center "Krasnoyarsk Science Center SB RAS", Krasnoyarsk, Russia.
authors.forename: Vasilisa V
authors.initials: VV
authors.lastname: Krasitskaya
authors.affiliation: Institute of Biophysics SB RAS, Federal Research Center "Krasnoyarsk Science Center SB RAS", Krasnoyarsk, Russia.
authors.forename: Ludmila P
authors.initials: LP
authors.lastname: Burakova
authors.affiliation: Siberian Federal University, Krasnoyarsk, Russia.
authors.forename: Anastasia A
authors.initials: AA
authors.lastname: Komarova
authors.affiliation: Institute of Biophysics SB RAS, Federal Research Center "Krasnoyarsk Science Center SB RAS", Krasnoyarsk, Russia.|Siberian Federal University, Krasnoyarsk, Russia.
authors.forename: Eugenia E
authors.initials: EE
authors.lastname: Bashmakova
authors.affiliation: Institute of Biophysics SB RAS, Federal Research Center "Krasnoyarsk Science Center SB RAS", Krasnoyarsk, Russia.|Siberian Federal University, Krasnoyarsk, Russia.
authors.forename: Ludmila A
authors.initials: LA
authors.lastname: Frank
mesh_terms: D002118:Calcium / Q000378:metabolism*; D003116:Color; D003545:Cysteine / Q000737:chemistry; D049449:Luminescence; D008164:Luminescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D020641:Polymorphism, Single Nucleotide; D011485:Protein Binding
chemical_list: D008164:Luminescent Proteins; C010497:obelin; D003545:Cysteine; D002118:Calcium