%0 Journal Article %A Zhaohua Zheng %A Syazwani I Amran %A Philip E Thompson %A Ian G Jennings %T Isoform Selective Inhibition of Phosphoinositide 3-kinase: Identification of a New Region of nNon-conserved Amino Acids Critical for p110α Inhibition. %D 2011 %R 10.1124/mol.111.072546 %J Molecular Pharmacology %P mol.111.072546 %X The combination of molecular modelling and x-ray crystallography has failed to yield a consensus model of the mechanism for selective binding of inhibitors to the phosphoinositide 3-kinase (PI3K) p110 α-isoform. Here we have used kinetic analysis to determine that the p110α selective inhibitor, PIK-75, is a competitive inhibitor with respect to a substrate, phosphatidylinositol (PI) in contrast to most other PI3K inhibitors which bind at or near the ATP site. Using sequence analysis and the existing crystal structures of inhibitor complexes with the p110γ and δ isoforms we have identified a new region of non-conserved amino acids (region 2) which was postulated to be involved in PIK-75 p110α selectivity. Analysis of region 2, using in vitro mutation of identified non-conserved amino acids to alanine, showed that S773 was a critical amino acid involved in PIK-75 binding with an 8-fold-increase in the IC-50 compared to wild-type. Kinetic analysis showed that, with respect to PI, the PIK-75 Ki for the isoform mutant S773D increased 64-fold as compared to wild type enzyme. In addition, a non-conservative amino acid, H855 from the previously identified region 1 of non-conserved amino acids was found to be involved in PIK-75 binding. These results show that these two regions of non-conserved amino acids that are close to the substrate binding site could be targeted in order to produce p110α isoform selective inhibitors. %U https://molpharm.aspetjournals.org/content/molpharm/early/2011/07/21/mol.111.072546.full.pdf