植物的生命周期和发育需要生物合成,沉积,细胞壁基质多糖的降解。不同细胞壁基质多糖的结构影响植物细胞的商业重要特性,包括增长,生物量不顺应性,器官脱落,和水果的保质期。这篇综述是对基质多糖糖基转移酶(GT)活性的全面总结,该活性已在异源GT蛋白表达后使用体外测定进行了验证。植物细胞壁(PCW)生物合成的GT主要是位于植物分泌系统的内质网和高尔基体的完整跨膜蛋白。这些酶在植物组织中的丰度较低,使得它们特别难以从天然植物膜中纯化出足以进行酶表征的数量。这对于研究不同GT的功能至关重要。在主要细胞壁基质聚糖的合成中的许多活性,包括果胶,木聚糖,木葡聚糖,甘露聚糖,混合键葡聚糖(MLGs),AGP蛋白聚糖的阿拉伯半乳聚糖成分已定位到特定基因和多基因家族。细胞壁GTs包括合成聚合物主链的那些,那些用延伸的糖基链延长侧分支的,以及将单个单糖连接添加到多糖主链和/或侧分支上的那些。已使用三种主要策略来鉴定编码合成细胞壁连接的GTs的基因:分析富集细胞壁生物合成活性的膜组分,研究细胞壁组成表型的突变遗传学方法,和来自测序植物基因组的推定GTs的组学指导鉴定。在这里,我们比较了用于生产的异源表达系统,净化,并研究PCWGTs的酶活性,重点是真核系统Nicotianabenthamiana,巴斯德毕赤酵母,和人胚胎肾(HEK293)细胞。我们讨论了GTs的酶学性质,包括动力学速率,多糖产品的链长,受体寡糖偏好,合成长链聚合物的伸长机理,以及GT复合物的形成。提出了基质多糖生物合成研究的未来方向。
The life cycle and development of plants requires the biosynthesis, deposition, and degradation of cell wall matrix polysaccharides. The structures of the diverse cell wall matrix polysaccharides influence commercially important properties of plant cells, including growth, biomass recalcitrance, organ abscission, and the shelf life of fruits. This
review is a comprehensive summary of the matrix polysaccharide
glycosyltransferase (GT) activities that have been verified using in vitro assays following heterologous GT protein expression. Plant cell wall (PCW) biosynthetic GTs are primarily integral transmembrane proteins localized to the endoplasmic reticulum and Golgi of the plant secretory system. The low abundance of these enzymes in plant tissues makes them particularly difficult to purify from native plant membranes in quantities sufficient for enzymatic characterization, which is essential to study the functions of the different GTs. Numerous activities in the synthesis of the major cell wall matrix glycans, including pectins, xylans, xyloglucan, mannans, mixed-linkage glucans (MLGs), and arabinogalactan components of AGP proteoglycans have been mapped to specific genes and multi-gene families. Cell wall GTs include those that synthesize the polymer backbones, those that elongate side branches with extended glycosyl chains, and those that add single monosaccharide linkages onto polysaccharide backbones and/or side branches. Three main strategies have been used to identify genes encoding GTs that synthesize cell wall linkages: analysis of membrane fractions enriched for cell wall biosynthetic activities, mutational genetics approaches investigating cell wall compositional phenotypes, and omics-directed identification of putative GTs from sequenced plant genomes. Here we compare the heterologous expression systems used to produce, purify, and study the enzyme activities of PCW GTs, with an emphasis on the eukaryotic systems Nicotiana benthamiana, Pichia pastoris, and human embryonic kidney (HEK293) cells. We discuss the enzymatic properties of GTs including kinetic rates, the chain lengths of polysaccharide products, acceptor oligosaccharide preferences, elongation mechanisms for the synthesis of long-chain polymers, and the formation of GT complexes. Future directions in the study of matrix polysaccharide biosynthesis are proposed.