Molecular Aspects of B-Glucan Biosynthesis in Higher Plants

;​你是说，我是说，我是说，我是说，我是说，我是说，我是说。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 ?lqz4 @ C， D， E， F， H， J， K， L， MF Microsoft Word 6.0文档MSWordDoc。6;​高等植物中callose (1,3)-(-linked glucan)的生物合成是由质膜结合糖基转移酶callose synthase (UDP-Glc: (1,3)-(-glucan synthase)催化的。胼胝质合酶活性在各种植物的分离膜组分中普遍存在，并且被假设为纤维素合酶的一种不受调节的形式。因此，为了了解细胞壁多糖生物生成和创面愈合调控的生化基础，有必要对多糖合酶（如胼胝质合酶）的多肽亚基及其编码基因进行鉴定和表征。红甜菜（Beta vulgaris L.）贮藏组织含有丰富而稳定的胼胝质合酶。纯化的胼胝质合成酶含有31、29和27 kDa多肽和一个弱染色的57 kDa蛋白。强疏水31和27 kDa多肽是主要内在蛋白（MIP）家族的质膜定位成员。本项目的假设是胼胝质合成酶是一种多聚体复合物，含有与跨膜蛋白密切相关的催化亚基，形成新生葡聚糖易位的通道。这个想法将通过三种方法进行测试。免疫和生化实验将查明胼胝质合成酶复合物组分负责糖基转移酶活性和聚合物易位。遗传学上，这些多肽将被克隆和测序，以确定它们的主要结构，在质膜内的取向，并确定调节基因表达的因素。功能上，重构实验将把特定的酶学和运输特性与单个多肽联系起来。总的来说，这些研究将把糖基转移和聚合物易位过程整合到一个单一的结构模型中。纤维素聚合物是世界上最丰富的天然可再生生物分子之一。生理上，来自植物细胞壁的纤维素聚合物提供了结构完整性和防止入侵植物病原体的保护。工业上，含木产品无处不在，纤维素是生产纸张、纺织品、食品、饲料、燃料和药品的成分。尽管这些材料具有重要的经济意义，但形成植物纤维素聚合物的生化和遗传机制尚不清楚。这项工作的目的是分离负责生产纤维素生物聚合物的基因和蛋白质，在高等植物中也称为-葡聚糖。(-葡聚糖合成复合物将从植物细胞外膜中提取并纯化。有了这些信息，免疫学和遗传学探针将被用来分离和测序编码-葡聚糖合成酶蛋白亚基的基因。证据表明，该复合物由一类亚基组成，一类亚基延长聚合物链，另一类亚基充当膜孔。这些孔为聚合物进入细胞壁提供了通道或孔。为了阐明这些膜通道的生化功能，将利用重组DNA技术进行表达研究。这个系统将提供对植物生长发育的了解。*** Oh +' 0 $ H l D H R:\WWUSER\TEMPLATE\NORMAL。点 沃瑟曼,布鲁斯·p·罗伯特·乌芬·雪莱·a·格雷夫斯（Bruce P. Robert Uffen，雪莱·a·格雷夫斯）@ S·m·m·a·格雷夫斯(雪莱·a·格雷夫斯Microsoft Word 6.0 2；​e = e j j j j j j j j 1 # % % % % j

; 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:\WWUSER\TEMPLATE\NORMAL.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