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Mutational Optimization Of The Coelenterazine-Dependent Luciferase From Renilla, Jongchan Woo, Albrecht G. Von Arnim Sep 2008

Mutational Optimization Of The Coelenterazine-Dependent Luciferase From Renilla, Jongchan Woo, Albrecht G. Von Arnim

Faculty Publications and Other Works -- Biochemistry, Cellular and Molecular Biology

Renilla luciferase (RLUC) is a popular reporter enzyme for gene expression and biosensor applications, but it is an unstable enzyme whose catalytic mechanism remains to be elucidated. We titrated that one RLUC molecule can turn over about one hundred molecules of coelenterazine substrate. Mutagenesis of active site residue Pro220 extended the half-life of photon emission, yielding brighter luminescence in E. coli. Random mutagenesis uncovered two new mutations that stabilized and increased photon emission in vivo and in vitro, while ameliorating substrate inhibition. Further amended with a previously identified mutation, a new triple mutant showed a threefold improved k …


Gene Alterations By Peroxisome Proliferator-Activated Receptor Gamma Agonists In Human Colorectal Cancer Cells, Maria Cekanova, X Li, J Yuan, K B. Kim, Seung J. Baek Apr 2008

Gene Alterations By Peroxisome Proliferator-Activated Receptor Gamma Agonists In Human Colorectal Cancer Cells, Maria Cekanova, X Li, J Yuan, K B. Kim, Seung J. Baek

Faculty Publications and Other Works -- Biochemistry, Cellular and Molecular Biology

The peroxisome proliferator-activated receptor gamma (PPARgamma) is a nuclear transcription factor that controls the genes involved in metabolism and carcinogenesis. In the present study, we examined the alteration of gene expression in HCT-116 human colorectal cancer cells by PPARgamma agonists: MCC-555 (5 microM), rosiglitazone (5 microM), and 15-deoxy-Delta12,14-prostaglandin J2 (1 microM). The long-oligo microarray data revealed a list of target genes commonly induced (307 genes) and repressed (32 genes) by tested PPARgamma agonists. These genes were analyzed by Onto-Express software and KEGG pathway analysis and revealed that PPARgamma agonists are involved in cell proliferation, focal adhesion, and several signaling pathways. …


Mutational Optimization Of The Coelenterazine-Dependent Luciferase From Renilla, Jongchan Woo, Albrecht G. Von Arnim Jan 2008

Mutational Optimization Of The Coelenterazine-Dependent Luciferase From Renilla, Jongchan Woo, Albrecht G. Von Arnim

Faculty Publications and Other Works -- Biochemistry, Cellular and Molecular Biology

Renilla luciferase (RLUC) is a popular reporter enzyme for gene expression and biosensor applications, but it is an unstable enzyme whose catalytic mechanism remains to be elucidated. We titrated that one RLUC molecule can turn over about one hundred molecules of coelenterazine substrate. Mutagenesis of active site residue Pro220 extended the half-life of photon emission, yielding brighter luminescence in E. coli. Random mutagenesis uncovered two new mutations that stabilized and increased photon emission in vivo and in vitro, while ameliorating substrate inhibition. Further amended with a previously identified mutation, a new triple mutant showed a threefold improved k …