Objective

Eukaryotic initiation factor 4A (eIF4A) is the ATP dependent helicase motor of the wider eIF4F complex that clears the 5’UTR of regulatory secondary structure elements such as G-quadruplexes. This allows a smooth landing pad for the 43S preinitiation complex to bind and subsequently scan for the translational start codon. The regulatory control exerted by eIF4A-mediated removal of G-quadruplex structures is highly associated with triple negative breast cancer, suggesting eIF4a as a therapeutic target. However, initial biochemical assay development in preparation for high throughput screening revealed an early product inhibition effect of ADP on the helicase activity. This study highlights the use of a Molecular Devices FLIPR Tetra to overcome this product inhibition by recording the very initial rate of activity in a high throughput assay format.