Biosynthesis of monoterpenes: Stereochemical implications of acyclic and monocyclic olefin formation by (+)- and (−)-pinene cyclases from sage

Abstract

(+)-Pinene cyclase from sage (Salvia officinalis) catalyzes the isomerization and cyclization of geranyl pyrophosphate to (+)-alpha-pinene and (+)-camphene, and to lesser amounts of (+)-limonene, myrcene, and terpinolene, whereas (-)-pinene cyclase from this tissue catalyzes the conversion of the acyclic precursor to (-)-alpha-pinene, (-)-beta-pinene, and (-)-camphene, and to lesser quantities of (-)-limonene, myrcene, and terpinolene. The bicyclic products of these enzymes (pinene and camphene) are derived via the cyclization of the cisoid, anti-endo-conformers of the bound, tertiary allylic intermediates (3R)-linalyl pyrophosphate [+)-pinene cyclase) and (3S)-linalyl pyrophosphate [-)-pinene cyclase). When challenged with either enantiomer of linalyl pyrophosphate or with neryl pyrophosphate (cis-isomer of geranyl pyrophosphate) as substrate, both pinene cyclases synthesize disproportionately high levels of acyclic olefins (myrcene and ocimene) and monocyclic olefins (limonene and terpinolene), compared with the product mixtures generated from the natural geranyl precursor. Resolution of the limonene derived from linalyl pyrophosphate and neryl pyrophosphate demonstrated that this monocyclic olefin was formed via conformational foldings in addition to the cisoid,anti-endo-pattern. These results indicate that the alternate substrates are ionized by the cyclases prior to their achieving the optimum orientation for bicyclization. In the case of geranyl pyrophosphate, a preassociation mechanism is suggested in which optimum folding of the terpenyl chain precedes the initial ionization step.