SGER: Synthetic Genes for the Design of Hydroxproline-Rich Glycomodules

SGER：用于设计富含羟脯氨酸的糖模块的合成基因

• 批准号: 9805960
• 负责人: Marcia Kieliszewski
• 金额: $ 5万
• 依托单位: Ohio University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-06-15 至 1999-11-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9805960&HistoricalAwards=false
• 关键词: SGER; Synthetic; Genes; Design; Hydroxproline

9805960 Kieliszewski Hydroxyproline-rich glycoproteins (HRGPs) are ubiquitous architectural components of growing plant cell walls, often accounting for 10-20% of their dry weight; polysaccharides account for the bulk. HRGPs are implicated in all aspects of plant growth and development including responses to stress. The HRGP superfamily contains three major groups representing a continuum of glycosylation and periodicity. They range from the repetitive proline-rich proteins that are lightly glycosylated and highly periodic, through the crosslinked extensins that are highly arabinosylated and periodic, to the arabinogalactan-proteins that are the most highly glycosylated and least periodic. The repetitive units are small, often only 4-6 residue glycosylated motifs viewed hypothetically as functional modules. The function of these unique plant glycoproteins remains largely unexplored. We propose to define their roles through functional analysis of single glycopeptide modules when expresssed as repetitive glycopolypeptides in tomato cultures. We shall insert synthetic genes encoding HRGP glycopeptide modules into two plant transformation vectors both derived from pBI121; one with a reporter gene, the other, for test for module function, lacking the reporter. We shall use these plasmids to transform cell cultures of tomato, which contain the plant prolylhydroxylases and glycosyltransferases required for module posttranslational modifications. Expression and isolation of these modules will enable us to test general and specific hypotheses for glycosylation and molecular function: 1. Glycosylation codes predicted by the Hyp-contiguity hypothesis specify precise saccharide addition to specific HRGP Hyp residues, namely: tetraarabinose to the most highly contiguous residues, arabinosylation decreasing with decreasing Hyp-contiguity; and polygalactosylation of repetitive noncontiguous Hyp residues that typify AGPs. 2. Carbohydrate-mediated alignment of extensin monomers is necessary for their orderly assembly and crosslinking roles in cell wall self-assembly.

富含羟脯氨酸的糖蛋白（HRGPs）是生长植物细胞壁中普遍存在的结构成分，通常占其干重的10-20%；多糖占大部分。hrgp参与植物生长发育的各个方面，包括对胁迫的反应。HRGP超家族包含三个主要基团，代表连续的糖基化和周期性。它们的范围从富含脯氨酸的重复蛋白（轻度糖基化且周期高），到交联延伸蛋白（高度阿拉伯糖基化且周期高），再到阿拉伯半乳糖蛋白（糖基化程度最高且周期最低）。重复单元很小，通常只有4-6个残基糖基化基序被认为是功能模块。这些独特的植物糖蛋白的功能在很大程度上仍未被探索。我们建议通过单个糖肽模块的功能分析来定义它们在番茄培养物中作为重复糖共肽表达时的作用。我们将编码HRGP糖肽模块的合成基因插入两个植物转化载体中，这两个载体都是从pBI121衍生的；一个带有报告基因，另一个用于测试模块功能，缺乏报告基因。我们将使用这些质粒转化番茄细胞培养物，其中含有模块翻译后修饰所需的植物脯氨酸羟化酶和糖基转移酶。这些模块的表达和分离将使我们能够测试糖基化和分子功能的一般和特定假设：通过高邻接假设预测的糖基化编码精确地指定了特定HRGP高邻接残基上的糖基化，即：四阿拉伯糖被添加到最高度邻接的残基上，阿拉伯糖基化随着低邻接程度的降低而降低；重复的不连续的Hyp残基的多半乳糖基化是典型的agp。2. 碳水化合物介导的延伸蛋白单体排列对于它们在细胞壁自组装中的有序组装和交联作用是必要的。