Poster: Secondary Metabolism

Abs # 711: Hydroxamic acid glucoside glucosidases of rye (Secale cereale)

Presenter:	Nikus, Jeanette , jeanette.nikus@sh.se
Additional Authors	Esen, Asim  (B)   Jonsson, Lisbeth MV (C)
Affiliations:	(A): Department of Plant Biology, Swedish University of Agricultural Science, Box 7080, SE-75007 Uppsala, Sweden (B): Department of Biology, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0406 (C): Natural Science Section, Södertörn University College, Box 4101, SE-14104 Huddinge, Sweden

We have investigated the presence and subcellular localization of b-glucosidases acting on hydroxamic acid glucosides in rye. The cyclic hydroxamic acids DIBOA and DIMBOA are secondary metabolites and occur in abundance as glucosides in maize, wheat and rye. They have been ascribed a role in plant defense. Upon tissue disruption the hydroxamic acid glucosides are cleaved by b-glucosidase to release toxic aglucones that are unstable and decompose to the reactive benzoxazolinones. In previous immunocytochemical studies we located rye b-glucosidase in plastids, cell wall and cytoplasm of coleoptiles, leaves and roots. The present study aims at characterizing the different b-glucosidases in order to analyze their possible function in the proposed Hx-glc/b-glucosidase defense system. b-Glucosidase cDNA was isolated from rye shoots, cloned and sequenced. Analysis of the precursor protein sequence with appropriate software indicated a N-terminal transit peptide of 49 amino acids for plastid targeting. The cDNA sequence corresponding to mature protein was expressed in E. coli and the biochemical properties of the recombinant protein were determined. The recombinant protein had the same physico-chemical properties as did native soluble protein isolated from rye. Both showed high activity with DIMBOA-glc or DIBOA-glc as substrate. In addition, an other b-glucosidase was partially purified from the cell wall fraction. The cell wall b-glucosidase differed from the plastid b-glucosidase in molecular weight, hydrophobic properties and substrate affinity but did exhibit activity with Hx-glc as substrate. b-Glucosidase activity was observed throughout the plant with the highest activities in the coleoptile and mesocotyle for both the plastid enzyme as well as the cell wall enzyme.