Actin filament- and Wiskott-Aldrich syndrome protein-binding sites on fructose-1,6-bisphosphate aldolase are functionally distinct from the active site.

The glycolytic enzyme fructose 1,6-(bis)phosphate aldolase (aldolase) is not only required for efficient utilization of glucose and fructose, but also for cytoskeletal functions like cytokinesis and cell motility. These differing roles are mediated by distinct and discrete binding interactions with aldolase's many binding partners, including actin filaments, Wiskott-Aldrich Syndrome protein (WASP), and Sorting Nexin 9 (SNX9). How these interactions are coordinated on the aldolase homotetramer of 160 kDa is unclear. In this study, the catalytic activity of wild-type aldolase is measured in the presence of actin filaments, and a WASP-derived peptide that binds to aldolase, or both. No appreciable changes in kcat or Km values are seen. Then, aldolase variants with substitutions targeting the tryptophan-binding pocket for WASP and SNX9 are created and perturbation of actin filament-, WASP peptide-, and SNX9 peptide-binding are assessed. Those that negatively impacted binding did not show an impact on aldolase catalysis. These results suggest that aldolase can engage in catalysis while simultaneously interacting with cytoskeletal machinery.