ACTIVATION OF POLLEN-TUBE CALLOSE SYNTHASE BY DETERGENTS - EVIDENCE FOR DIFFERENT MECHANISMS OF ACTION

In pollen tubes of Nicotiana alata, a membrane-bound, Ca2+ independent callose synthase (CalS) is responsible for the biosynthesis of the (1 ,3)-beta-glucan backbone of callose, the main cell wall component. Dig itonin increases CalS activity 3-to 4-fold over a wide range of concen trations, increasing the maximum initial velocity without altering the Michaelis constant for UDP-glucose. The CalS activity that requires d igitonin for assay (the latent CalS activity) is not inhibited bythe m embrane-impermeant, active-site-directed reagent UDP-pyridoxal when th e reaction is conducted in the absence of digitonin. This is consisten t with digitonin increasing CalS activity bythe permeabilization of me mbrane vesicles. A second group of detergents, including olamidopropyl )dimethylammonio]-1-propane-sulfonate (CHAPS), Zwittergent 3-16, and 1 -alpha-lysolecithin, activate pollen tube CalS 10-to 15-fold, but only over a narrow range of concentrations just below their respective cri tical micellar concentrations. This activation could not be attributed to any particular chemical feature of these detergents. CHAPS increas es maximum initial velocity and decreases the Michaelis constant for U DP-glucose and activates CalS even in the presence of permeabilizing c oncentrations of digitonin. Inhibition studies with UDP-pyridoxal indi cate that activation by CHAPS occurs by recruitment of previously inac tive CalS molecules to the pool of active enzyme. The activation of po llen tube CalS by these detergents therefore resembles activation of t he enzyme by trypsin.