KRAS Isoform B Coupled Nucleotide Exchange (TR-FRET) Assay Kit

The KRAS Isoform B Coupled Nucleotide Exchange TR-FRET Assay Kit is a TR-FRET (Time-Resolved Fluorescence Resonance Energy Transfer) assay kit designed for screening and profiling of KRAS Isoform B antagonists/inhibitors. It monitors the binding of an effector protein such as the Ras binding domain of RAF1, (RBD-cRaf) to KRAS. The KRAS Isoform B Coupled Nucleotide Exchange TR-FRET Assay Kit comes in a convenient 384-well format, with enough purified recombinantGDP-loaded KRAS Isoform B, GTP, exchange factor SOS1 (son of sevenless homolog 1) (amino acids 564-1049), an effector protein RBD-cRAF (amino acids 50-140), assay buffer and additives for 400 reactions.Figure 1: Illustration of the KRAS Isoform B Coupled Nucleotide Exchange TR-FRET Assay Kit principle.A sample containing GDP-loaded KRAS Isoform B is incubated with SOS1 and GTP, allowing nucleotide exchange. GDP-loaded KRAS Isoform B is in an inactive state and does not interact with the Ras-binding domain (RBD) of cRAF. SOS1 assists in the release of GDP from KRAS so that GTP can occupy the nucleotide binding pocket. This results in a conformational change in KRAS that permits its binding to RBD-cRAF. RBD-cRAF is then added to the reaction. After incubation, acceptor and donor dyes are added. The TR-FRET signal generated can be measured using a fluorescence plate reader. The signal intensity is proportional to the presence of GTP-bound KRAS.Need us to run inhibitor screens or profile your compounds that block KRAS Isoform B exchange? Check out ourKRAS Screening & Profiling Services.Need us to run inhibitor screens or profile your compounds that block KRAS Isoform B exchange? Check out ourKRAS Screening & Profiling Services.