Membrane association and some characteristics of the ethylene forming enzyme from etiolated pea seedlings

Abstract

When freshly prepared homogenates of etiolated pea ( Pisum sativum L. cv Calvedon) subhook segments were fractionated by high speed centrifugation, the enzyme catalyzing the conversion of 1-aminocyclopropane-l-carboxylic acid (ACC) to ethylene was found associated with the particulate fraction. However, on aging the homogenates at 5°C prior to fractionation, 50 to 75% of the enzyme activity partitioned into the soluble fraction; this solubilization led to a highly activated enzyme form. Both the particulate and soluble enzyme exhibited non-linear substrate saturation kinetics and were inhibited to similar extents by ascorbic acid, EDTA, CoCl 2 and limiting oxygen. However, they differed in their response to incubation with n-propylgallate, dithiothreitol, CaCl 2 and 100% oxygen. Calcium stimulated only the particulate form and increased both the ‘low K m ’ for ACC from 2.99 to 5.58 mM and apparent Vmax from 88 to 285 nl/mg protein/h.