The cover picture shows a TR-FRET-based functional assay for screening small-molecule modulators of co-activator-associated arginine methyltransferase 1 (CARM1), based on its methylation of PABP1. The formation of the ternary complex (Me-GFP-PABP1, Me-PABP1 Ab, and Tb-2nd Ab) enables time-resolved detection of the long-lifetime terbium (Tb)-to-GFP FRET signal. Light at 340 nm is used to excite Tb, which emits at 495 nm. The proximity of the donor Tb to the acceptor GFP results in energy transfer from Tb to GFP, and this induces the specific FRET signal at 520 nm. On p. 827 ff., W. Xu et al. describe the development and optimization of the homogenous Me-GFP-PABP1-based TR-FRET assay for monitoring the expression and enzymatic activity of CARM1 in MCF7 breast cancer cells. The assay has been optimized for high-throughput screening to identify CARM1 allosteric activators. More importantly, this methodology has the potential to be extended to other protein arginine methyltransferases through the development of appropriate substrate–GFP fusion proteins and methylation-site-specific antibodies.

Cover Picture: A TR-FRET-Based Functional Assay for Screening Activators of CARM1 (ChemBioChem 7/2013)

SBARDELLA, Gianluca;
2013

Abstract

The cover picture shows a TR-FRET-based functional assay for screening small-molecule modulators of co-activator-associated arginine methyltransferase 1 (CARM1), based on its methylation of PABP1. The formation of the ternary complex (Me-GFP-PABP1, Me-PABP1 Ab, and Tb-2nd Ab) enables time-resolved detection of the long-lifetime terbium (Tb)-to-GFP FRET signal. Light at 340 nm is used to excite Tb, which emits at 495 nm. The proximity of the donor Tb to the acceptor GFP results in energy transfer from Tb to GFP, and this induces the specific FRET signal at 520 nm. On p. 827 ff., W. Xu et al. describe the development and optimization of the homogenous Me-GFP-PABP1-based TR-FRET assay for monitoring the expression and enzymatic activity of CARM1 in MCF7 breast cancer cells. The assay has been optimized for high-throughput screening to identify CARM1 allosteric activators. More importantly, this methodology has the potential to be extended to other protein arginine methyltransferases through the development of appropriate substrate–GFP fusion proteins and methylation-site-specific antibodies.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11386/4656355
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