Trehalase Activity in Arabidopsis thaliana Optimized for 96-well Plates

Abstract

Trehalose is a nonreducing disaccharide. It is a common sugar in bacteria, fungi and yeast, where it functions as a carbon source and stress protectant. In contrast, plants, although encoding large trehalose biosynthesis gene families, contain only small amounts of trehalose. The intermediate compound of trehalose, trehalose-6-phosphate (T6P), is a signaling molecule in plants, regulating metabolism, growth, and development. Most plants contain only a single trehalase, the enzyme that specifically hydrolyzes trehalose into two glucose molecules. High trehalase activity has been suggested to be part of the defense mechanism in plants hosting mycorrhizal fungi, rhizobia, and the plant pathogen Plasmodiophora brassica. Recently, it was shown in Arabidopsis thaliana that high trehalase activity is associated with an increase in drought stress tolerance and that trehalase fulfills an important role in stomatal regulation. Here we describe a protocol for measuring trehalase activity in Arabidopsis tissues, optimized for 96-well plates. Dialyzed protein extracts will be incubated with trehalose, followed by the quantitation of the released glucose using glucose oxidase-peroxidase.