Enzymic synthesis of 1-O-(indol-3-ylacetyl)-β-d-glucose. Purification of the enzyme from Zea mays, and preparation of antibodies to the enzyme

Abstract

The enzyme indol-3-ylacetylglucose synthase (UDP-glucose:indol-3-ylacetate beta-D-glucosyltransferase) catalyses the reaction: [formula: see text] This is the first step in the series of reactions leading to the indol-3-ylacetic acid conjugates found in maize. Previous attempts to purify this enzyme from the liquid endosperm of kernels of Zea mays (sweet corn) were not entirely successful owing to the lability of partially purified preparations during column chromatography. Thus this enzyme has not previously been purified to homogeneity. During the present study it was found that retention of enzyme activity required the combined presence of glycerol and dithiothreitol. Adding these requirements permitted purification of the enzyme to homogeneity with retention of catalytic activity. These purified preparations were used for preparation of rabbit polyclonal antibodies to the enzyme. Antibodies to the Zea mays endosperm enzyme cross-react with the enzyme from Zea mays vegetative tissues and with an enzyme from the liquid endosperm of oak acorns (Quercus sp). In this paper we report a simplified purification procedure adaptable to the preparation of milligram amounts of the enzyme.