Engineering of Decameric D-fructose-6-phosphate Aldolase A by Combinatorial Modulation of Inter- and Intra-subunit Interactions

Combinatorial modulation of inter- and intra-subunit interactions on decameric D-fructose-6-phosphate aldolase A (FSAA) generated a triple-site variant I31T/Q59T/I195Q FSAA with 27 to 278-fold activity improvement towards target heteroaromatic aldehydes. X-ray crystallographic data and molecular dynamics simulations ascribed the enhanced activity to pronounced flexibility of interface region between subunits, expanded substrate entrance and binding pocket, and enhanced proton transfer, unambiguously demonstrating the efficiency of this strategy for engineering multimeric enzymes.