Phylogenetic variation in glycosidases and glycanases acting on plant cell wall polysaccharides, and the detection of transglycosidase and trans-β-xylanase activities
Phylogenetic variation in glycosidases and glycanases acting on plant cell wall polysaccharides, and the detection of transglycosidase and trans-β-xylanase activities
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DOI：
10.1111/j.1365-313x.2011.04625.x
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发表时间：
2011-08-01
期刊：
影响因子：
7.2
通讯作者：
Fry, Stephen C.
中科院分区：
文献类型：
--
作者：
Frankova, Lenka;Fry, Stephen C.
细胞壁多糖的化学性质在系统发育上是不同的，这表明细胞壁酶需要变异。虽然植物拥有许多被认为作用于壁碳水化合物的酶的基因，但编码蛋白质的活性通常仍然是推测性的。为了探索可证明的酶活性的系统发育差异，我们用三种萃取剂从57种快速生长的植物器官中提取了蛋白质，并分析了它们对六种低聚糖的作用能力，这些低聚糖“模拟”了选定的细胞壁多糖。根据反应产物，我们成功地区分了外水解酶和内水解酶，并发现筛选的所有水解酶的分类差异很大：β - d -木糖苷酶、内do-(1 -> - 4)- β - d -木聚糖酶、β - d -甘露糖苷酶、内do-(1 -> - 4)- β - d -甘露聚糖酶、α - d -木糖苷酶、β - d -半乳糖苷酶、α - l -阿拉伯糖苷酶和α - l -聚焦酶。结果作为GHATAbase，一个可搜索的Excel格式的汇编，也为选择作用于壁碳水化合物的酶的丰富来源提供了一个汇编。其中四种水解酶伴随着，有时超过，转糖基化酶的活性，产生比底物大的产物。例如，在对(1 - bbbb4)- β - d -木糖己糖（Xyl(6)）进行β -木糖苷酶测定时，Marchantia， Selaginella和Equisetum提取物的游离木糖可以忽略，但大约等量的Xyl(5)和Xyl(7)，表明反式- β -木糖苷酶活性，洋葱，谷物，豆类和油菜中也有。Xyl(9)的产率经常超过Xyl(7-8)，说明β -木聚糖酶伴有内转糖基化酶活性，这里称为反式β -木聚糖酶，催化2Xyl(6) -> Xyl(3) + Xyl(9)的反应。类似的证据还表明，反式- α -木糖苷酶、反式- α -阿拉伯糖糖苷酶和反式- α -阿拉伯糖酶的活性作用于木葡聚糖低聚糖和(1 - bbbb5)- α - l -阿拉伯糖低聚糖。综上所述，不同植物作用于壁碳水化合物的可提取酶存在显著差异，反映了壁多糖组成的差异。除糖苷酶和糖聚糖酶活性外，还检测到5种新的转糖基化酶活性。我们认为这些活动在墙矩阵的组装和重组中起作用。
Wall polysaccharide chemistry varies phylogenetically, suggesting a need for variation in wall enzymes. Although plants possess the genes for numerous putative enzymes acting on wall carbohydrates, the activities of the encoded proteins often remain conjectural. To explore phylogenetic differences in demonstrable enzyme activities, we extracted proteins from 57 rapidly growing plant organs with three extractants, and assayed their ability to act on six oligosaccharides 'modelling' selected cell-wall polysaccharides. Based on reaction products, we successfully distinguished exo- and endo-hydrolases and found high taxonomic variation in all hydrolases screened: beta-D-xylosidase, endo-(1 -> 4)-beta-D-xylanase, beta-D-mannosidase, endo-(1 -> 4)-beta-D-mannanase, alpha-D-xylosidase, beta-D-galactosidase, alpha-L-arabinosidase and alpha-L-fucosidase. The results, as GHATAbase, a searchable compendium in Excel format, also provide a compilation for selecting rich sources of enzymes acting on wall carbohydrates. Four of the hydrolases were accompanied, sometimes exceeded, by transglycosylase activities, generating products larger than the substrate. For example, during beta-xylosidase assays on (1 -> 4)-beta-D-xylohexaose (Xyl(6)), Marchantia, Selaginella and Equisetum extracts gave negligible free xylose but approximately equimolar Xyl(5) and Xyl(7), indicating trans-beta-xylosidase activity, also found in onion, cereals, legumes and rape. The yield of Xyl(9) often exceeded that of Xyl(7-8), indicating that beta-xylanase was accompanied by an endotransglycosylase activity, here called trans-beta-xylanase, catalysing the reaction 2Xyl(6) -> Xyl(3) + Xyl(9). Similar evidence also revealed trans-alpha-xylosidase, trans-alpha-arabinosidase and trans-alpha-arabinanase activities acting on xyloglucan oligosaccharides and (1 -> 5)-alpha-L-arabino-oligosaccharides. In conclusion, diverse plants differ dramatically in extractable enzymes acting on wall carbohydrate, reflecting differences in wall polysaccharide composition. Besides glycosidase and glycanase activities, five new transglycosylase activities were detected. We propose that such activities function in the assembly and re-structuring of the wall matrix.