Hydrolysis and Isomerization of trans,trans‐Farnesyl Diphosphate by Andrographis Tissue‐Culture Enzymes

Abstract

1. Incubation of (3R,5S)-[5-³H₁]mevalonate + (3RS)-[2-¹⁴C]mevalonate with Andrographis cell-free extract leads to trans,trans-farnesol and cis,trans-farnesol which both totally retain tritium. 2. This conflicts with our previous results which predict one third tritium loss in the cis,trans-farnesol. Inversion at C-1 during hydrolysis of trans,trans-farnesyl diphosphate to trans,trans-farnesol could explain this anomaly. 3. (1S)-trans,trans-[1-³H₁]Farnesyl diphosphate and phosphate and (1R)-trans,trans-[1-³H₁]-farnesyl diphosphate and phosphate, all prepared chemically, were hydrolysed with Andrographis phosphatase, and alkaline phosphatase and hydrogenolysed with lithium aluminium hydride and the product alcohols exchanged with liver alcohol hydrogenase. 4. Both Andrographis phosphatase and alkaline phosphatase hydrolyse trans,trans-farnesyl diphosphate and trans,trans-farnesyl phosphate with retention. 5. Hydrolysis of trans,trans-[1-¹⁸O]farnesyl diphosphate in H₂¹⁸O with both phosphatases supports P-O fission. 6. The C-1 configuration in (1S)-trans,trans-[1-³H₁]farnesyl diphosphate and phosphate and (1R)-trans,trans-[1-³H₁]farnesyl diphosphate and phosphate is progressively racemised in 0.01 M NH₄OH/MeOH (1/9) at –20°C.