Triosephosphate Isomerase: Difference between revisions

The residues in the flexible loop are highly conserved among a variety of TIM enzymes from different organisms as an indication of their functional significance. Four residues in the flexible loop were deleted by site-directed mutagenesis to test the functional role of the loop region in catalysis. The deletion mutant was found to have a severe defect in kcat despite a modest defect in Km indicating a primary role for the loop region. Furthermore, a significant increase in the level of methylglyoxal was observed for the deletion mutant compared to the wild type enzyme. Methylglyoxal is a toxic by-product of the reaction, which builds up when the enediol intermediate is released from the active site. Thus, the flexible loop also functions to hold the intermediate in the active site so catalysis can proceed to the final product.