title: Novel functional Renilla luciferase mutant provides long-term serum stability and high luminescence activity.
abstract: Fluorescent and luminescent chemical probes are essential in recent medical diagnostics. However, the use of these probes in vivo has raised concerns due to their low sensitivity, background signal interference, and non-biocompatibility. Therefore, biological chromophores have received much attention as new alternatives. In particular, luciferase, a class of oxidative enzyme with bioluminescence, has emerged as a promising fluorophore due to its improved biocompatibility. However, the enzyme usually possesses weaker luminescence and stability relative to its chemically-based competitors. Here, we report a novel functional mutant luciferase with both enhanced luminescence and long-term serum stability. For the preparation of the modified Renilla luciferase, a new bacterial subcloning design was established. The luciferase coding DNA sequence was redesigned so that mutant luciferase could be easily expressed in an Escherichia coli system. The mutant Renilla luciferase, which we called "m-Rluc," demonstrated characteristic enzymatic functions and showed a 5.6-fold increase in luminescence activity. In addition, the enzyme's physiological stability remained >80% for more than 5days, in contrast to conventional luciferase, termed "hrluc," which disappeared within a few hours. We suggest that this novel biological luciferase probe may be a great tool for both in vitro and in vivo medical diagnostics.
authors: School of Chemical Engineering, Sungkyunkwan University, Suwon, Gyeonggi-do 440-746, South Korea.
authors: Woo Chul
authors: WC
authors: Song
authors: Hye-Jin
authors: HJ
authors: Sung
authors: Kyung Soo
authors: KS
authors: Park
authors: Jeong-Woo
authors: JW
authors: Choi
authors: Je-Yeol
authors: JY
authors: Cho
authors: Soong Ho
authors: SH
authors: Um
mesh_terms: D000818:Animals; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D008156:Luciferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D049449:Luminescence; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D048790:Renilla / Q000201:enzymology* / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D008156:Luciferases
keywords: Cloning; Diagnostics; Functional activity; Luciferase; Mutation