; R o o t E n t r y F Z?lqZ C o m p O b j b W o r d D o c u m e n t O b j e c t P o o l Z?lqZ Z?lqZ 4 @ C D E F G H I J K L M F Microsoft Word 6.0 Document MSWordDoc Word.Document.6 ; 9507266 Wasserman The biosynthesis of callose (1,3)-(-linked glucan) in higher plants is catalyzed by the plasma membrane-bound glycosyl transferase callose synthase (UDP-Glc: (1,3)-(-glucan synthase). Callose synthase activity is ubiquitous in isolated membrane fractions from a variety of plants, and has been hypothesized to be a deregulated form of cellulose synthase. Therefore, to understand the biochemical basis for the regulation of cell wall polysaccharide biogenesis and wound healing, it is necessary to identify and characterize the polypeptide subunits of polysaccharide synthases, like callose synthase, and the genes which encode them. Red beet (Beta vulgaris L.) storage tissue contains an abundant and stable callose synthase. Purified callose synthase contains 31, 29, and 27 kDa polypeptides and a weakly-staining 57 kDa protein. Strongly hydrophobic 31 and 27 kDa polypeptides are plasma membrane-localized members of the major intrinsic protein (MIP) family. The hypothesis of this project is that callose synthase is a multimeric complex containing a catalytic subunit closely associated with transmembrane proteins, forming a channel through which nascent glucans are translocated. The idea will be tested using three approaches. Immunological and biochemical experiments will pinpoint callose synthase complex components responsible for glycosyl transferase activity and polymer translocation. Genetically, these polypeptides will be cloned and sequenced to determine their primary structure, orientation within the plasma membrane, and to identify factors regulating gene expression. Functionally, reconstitution experiments will correlate specific enzymological and transport properties with individual polypeptides. Collectively, these studies will integrate the processes of glycosyl transfer and polymer translocation into a single structural model. %%% Cellulosic polymers are among the world s most abundant naturally-occurring, renewable biomolecules. Physiologically, cellulose polymers from plant cell walls provide structural integrity and protection against invading plant pathogens. Industrially, wood-containing products are ubiquitous, and cellulosics are components for the production of paper, textiles, foods, feeds, fuels, and pharmaceuticals. Despite the economic importance of these materials, the biochemical and genetic mechanisms are unclear which form plant cellulose polymers. The objective of this work is to isolate the genes and proteins responsible for the production of cellulosic biopolymers, also called (-glucans in higher plants. (-Glucan synthetic complexes will be extracted from the outer membrane of plant cells and purified. With this information, immunological and genetic probes will be made to isolate and sequence genes encoding protein subunits of (-glucan synthases. Evidence indicates that the complex consists of one class of subunits which elongate polymer chains, and a second class which serve as membrane pores. These pores provide a conduit, or aperture, for entry of the polymers into the cell wall. To elucidate the biochemical functions of these membrane channels, expression studies will be done with recombinant DNA techniques. This system will provide understanding of plant growth and development. *** Oh +' 0 $ H l D h R:WWUSERTEMPLATENORMAL.DOT Wasserman, Bruce P. Robert Uffen Shelley A. Graves @ S u m m a r y I n f o r m a t i o n ( B K @ @ ZqZ @ F # Microsoft Word 6.0 2 ; e = e j j j j j j j 1 # % % % % J
