Sesquiterpene alpha-farnesene synthase: Partial purification, characterization, and activity in relation to superficial scald development in apples

To decipher the relation between alpha-farnesene metabolism and the develop ment of superficial scald in apples, trans,trans-alpha-farnesene synthase, the enzyme that catalyzes the conversion of farnesyl pyrophosphate to alpha -farnesene, was partially purified from skin tissue of 'Delicious' apples ( Malus xdomestica Borkh.) and characterized. Total and specific activities o f the enzyme were higher in the cytosolic fraction than in membrane fractio ns. alpha-Farnesene synthase was purified 70-fold from the cytosolic fracti on by ion exchange chromatography and gel permeation, and the native molecu lar weight was estimated to be 108,000, The enzyme had optimal activity at a pH of 5.6 and absolutely required a divalent metal ion such as Mg2+ or Mn 2+ for activity. It exhibited allosteric kinetics, S-(0.5) for farnesyl pyr ophosphate being 84 +/- 18 mu mol.L-1, and a Hill coefficient (n(H)) of 2.9 , indicating the number of subunits to be two or three. Enzyme activity was highest between 10 and 20 degrees C, while 50% of the maximal activity was retained at 0 degrees C. In vivo alpha-farnesene synthase activity was min imal at harvest, then increased rapidly during 16 weeks storage in air at 0 degrees C, and decreased during further:storage. Activity of alpha-farnese ne synthase, alpha-farnesene content, and conjugated triene alcohol (the pu tative scald-causing oxidation product of alpha-farnesene) content in skin tissue were not correlated to the inherent nature of scald susceptibility o r resistance in 11 apple cultivars tested.