tailieunhanh - Báo cáo khoa học: The three desulfoglucosinolate sulfotransferase proteins in Arabidopsis have different substrate specificities and are differentially expressed

Sulfotransferases (SOTs) catalyse the transfer of a sulfate group from 3¢-phosphoadenosine 5¢-phosphosulfate (PAPS) to an appropriate hydroxy group of various substrates with the parallel formation of 3¢-phospho-adenosine 5¢-phosphate. In Arabidopsis thaliana, 18 SOT proteins (AtSOT) have been identified. | iFEBS Journal The three desulfoglucosinolate sulfotransferase proteins in Arabidopsis have different substrate specificities and are differentially expressed Marion Klein1 Michael Reichelt2 Jonathan Gershenzon2 and Jutta Papenbrock1 1 Institute for Botany University of Hannover Germany 2 Max Planck Institute for ChemicalEcology Jena Germany Keywords glucosinolates 3 -phosphoadenosine 5 -phosphosulfate PAPS subcellular localization substrate specificity sulfotransferase Correspondence J. Papenbrock Institute for Botany University of Hannover Herrenhauserstr. 2 D-30419 Hannover Germany Fax 49 511762 3992 Tel 49 511762 3788 E-mail Received 26 August 2005 revised 26 October 2005 accepted 3 November 2005 doi Sulfotransferases SOTs catalyse the transfer of a sulfate group from 3 -phosphoadenosine 5 -phosphosulfate PAPS to an appropriate hydroxy group of various substrates with the parallel formation of 3 -phospho-adenosine 5 -phosphate. In Arabidopsis thaliana 18 SOT proteins AtSOT have been identified. Three of them AtSOT16 AtSOT17 and AtSOT18 catalyse the sulfation of desulfoglucosinolates. The proteins were expressed in Escherichia coli purified by affinity chromatography and used for enzyme kinetic studies. By establishing two types of enzyme assay using both 35S-labelled and unlabelled PAPS separation of the products by HPLC and detection of the products by monitoring radioactivity or UV absorption the substrate specificities of the three AtSOT proteins were determined. They show different maximum velocities with several desulfo-glucosinolates as substrates and differ in their affinity for desulfobenzylglu-cosinolate and PAPS. The sequences encoding AtSOT18 were amplified from Arabidopsis ecotypes C24 and Col0 the two expressed proteins differ in two out of 350 amino acids. These amino-acid variations led to different substrate specificities. Exchange of one of the two amino acids in AtSOT18 .