title: Cation-driven Optical Properties of Artificial Luciferases.
abstract: The present study demonstrates cation-driven optical properties of artificial luciferases (ALucs) from copepod luciferases, as an optical readout for bioanalysis. An assignment of the supersecondary structure code (SSC) of ALucs revealed that ALucs carry a helix-loop-helix structure, which appears at the same sites of the EF-hands of typical Ca(2+)-binding proteins. A mutagenesis study shows that the EF-hand-like structure is a pivotal site for enzymatic activity. The effects of 20 kinds of mono- and multivalent cations on ALuc activities were estimated with column-purified ALuc16. High pH values boost the ALuc activities with both the native coelenterazine and an analog called 6-pi-OH-CTZ. Multivalent cations, Ca(II), Mg(II), and Cr(VI), elevate and prolong the ALuc activities, whereas Co(II), Cu(II) and Pb(II) greatly hamper the ALuc activities. Ca(II) greatly prolongs the optical intensities, suggesting a contribution to the structural robustness of ALucs. The inhibitory effect of multivalent cations on the ALuc activities was utilized for creating dose-response curves. The intrinsic cation-driven selectivity and optical intensity of ALucs enable researchers to constitute de novo biosensors for multivalent cations.
authors.affiliation: National Institute of Advanced Industrial Science and Technology (AIST), AIST Tsukuba West.
authors.forename: Sung Bae
authors.initials: SB
authors.lastname: Kim
authors.forename: Simon
authors.initials: S
authors.lastname: Miller
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Suzuki
authors.forename: Toshiya
authors.initials: T
authors.lastname: Senda
authors.forename: Ryo
authors.initials: R
authors.lastname: Nishihara
authors.forename: Koji
authors.initials: K
authors.lastname: Suzuki
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002412:Cations / Q000494:pharmacology; D033342:Copepoda / Q000201:enzymology*; D020832:EF Hand Motifs; D004795:Enzyme Stability / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D008156:Luciferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008163:Luminescent Measurements; D008958:Models, Molecular; D008969:Molecular Sequence Data; D055620:Optical Phenomena; D011522:Protons
chemical_list: D002412:Cations; D011522:Protons; D008156:Luciferases