Trypanosoma brucei prostaglandin F 2␣ synthase is an aldo-ketoreductase that catalyzes the reduction of prostaglandin H 2 to PGF 2␣ in addition to that of 9,10-phenanthrenequinone. We report the crystal structure of TbPGFS⅐NADP protonation through a water molecule, while exerting an electrostatic repulsion against His 110 that maintains the spatial arrangement which allows the formation of a hydrogen bond between His 110 and C 11 carbonyl of PGH 2 . We also show that Tyr 52 acts as the general acid catalyst for 9,10-PQ reduction, and thus we not only elucidate the catalytic mechanism of a PGH 2 reductase but also provide an insight into the catalytic specificity of AKRs.