Stereospecificity of (+)-pinoresinol and (+)-lariciresinol reductases from Forsythia intermedia

Pinoresinol/lariciresinol reductase catalyzes the first known example of a highly unusual benzylic ether reduction in plants; its mechanism of hydride transfer is described. The enzyme was found in Forsythia intermedia and catalyzes the presumed regulatory branch-points in the pathway leading to benzylaryltetrahydrofuran, dibenzylbutane, dibenzylbutyrolactone, and aryltetrahydronaphthalene lignans. Using [7,7′-²H₂]-pinoresinol and [7,7′-²H₃]lariciresinol as substrates, the hydride transfers of the highly unusual reductase were demonstrated to be completely stereospecific (>99%). The incoming hydrides were found to take up the pro-R position at C-7′ (and/or C-7) in lariciresinol and secoisolariciresinol, thereby eliminating the possibility of random hydride delivery to a planar quinone methide intermediate. As might be expected, the mode of hydride abstraction from NADPH was also stereospecific: using [4A-³H] and [4S-³H]NADPH, it was found that only the 4 pro-R hydrogen was abstracted for enzymatic hydride transfer.