Directed Molecular Evolution Reveals Gaussia Luciferase Variants with Enhanced Light Output Stability

被引：35
作者：
Degeling, M. Hannah [1 ,2 ,3 ]
Bovenberg, M. Sarah S. [1 ,2 ,3 ]
Lewandrowski, Grant K. [1 ,2 ]
de Gooijer, Mark C. [1 ,2 ,4 ]
Vleggeert-Lankamp, Carmen L. A. [3 ]
Tannous, Marie [5 ]
Maguire, Casey A. [1 ,2 ]
Tannous, Bakhos A. [1 ,2 ]
机构：
[1] Massachusetts Gen Hosp, Ctr Neurosci, Expt Therapeut & Mol Imaging Lab, Dept Neurol, Charlestown, MA 02129 USA
[2] Harvard Univ, Sch Med, Program Neurosci, Boston, MA 02114 USA
[3] Leiden Univ, Med Ctr, Dept Neurosurg, Leiden, Netherlands
[4] Canc Ctr Amsterdam, VU Med Ctr, Dept Neurosurg, Neurooncol Res Grp, NL-1007 MB Amsterdam, Netherlands
[5] Notre Dame Univ, Fac Nat & Appl Sci, Barsa, Lebanon
来源：
ANALYTICAL CHEMISTRY | 2013年 / 85卷 / 05期
关键词：
REPORTER GENE-EXPRESSION; BIOLUMINESCENCE;
D O I：
10.1021/ac4003134
中图分类号：
O65 [分析化学];
学科分类号：
070302 ; 081704 ;
摘要：
Gaussia Luciferase (Gluc) has proven to be a powerful mammalian cell reporter for monitoring numerous biological processes in immunology, virology, oncology, and neuroscience. Current limitations of Gluc as a reporter include its emission of blue light, which is absorbed by mammalian tissues, limiting its use in vivo, and a flash-type bioluminescence reaction, making it unsuited for high-throughput applications. To overcome these limitations, a library of Gluc variants was generated using directed molecular evolution and screened for relative light: output, a shift in emission spectrum, and glow-type light emission kinetics. Several variants with a 10-15 nm shift in their light emission peak were found. Further, a Gluc variant that catalyzes a glow-type bioluminescence reaction, suited for high-throughput applications, was also identified. These results indicate that molecular evolution could be used to modulate Gluc bioluminescence reaction characteristics.
引用
收藏
页码：3006 / 3012
页数：7
相关论文
共 23 条
[21]   Codon-optimized Gaussia luciferase cDNA for mammalian gene expression in culture and in vivo [J].
Tannous, BA ;
Kim, DE ;
Fernandez, JL ;
Weissleder, R ;
Breakefield, XO .
MOLECULAR THERAPY, 2005, 11 (03) :435-443
[22]   Imaging in the era of molecular oncology [J].
Weissleder, Ralph ;
Pittet, Mikael J. .
NATURE, 2008, 452 (7187) :580-589
[23]  
Wurdinger T, 2008, NAT METHODS, V5, P171, DOI [10.1038/nmeth.1177, 10.1038/NMETH.1177]
123