Molecular characterisation of a xyloglucan oligosaccharide-acting α-D-xylosidase from nasturtium (Tropaeolum majus L.) cotyledons that resembles plant 'apoplastic' α-D-glucosidases
Molecular characterisation of a xyloglucan oligosaccharide-acting α-D-xylosidase from nasturtium (Tropaeolum majus L.) cotyledons that resembles plant 'apoplastic' α-D-glucosidases
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DOI：
10.1007/s004250100631
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发表时间：
2002-01-01
期刊：
影响因子：
4.3
通讯作者：
Reid, JSG
中科院分区：
文献类型：
--
作者：
Crombie, HJ;Chengappa, S;Reid, JSG
本文报道了从发芽的水仙(Tropaeolum Majus L.)子叶中分离、测序和分析与木葡聚糖动员相关的α-D-木糖苷酶的基因。种子。翻译的开放阅读框(含终止密码子)为2,808个碱基，由935个氨基酸组成。它包括由蛋白质内切酶消化获得的11个肽的序列，以及一个假定的疏水信号序列，该序列是通过质膜定向的蛋白质的特征。推测的翻译蛋白的相对分子质量明显高于十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法测定的相对分子质量，提示翻译后修饰。该蛋白序列与拟南芥的α-木糖苷酶序列高度同源(896个氨基酸以上的同源性为76.0%)，并与多种α-葡萄糖苷酶有同源性，特别是与植物细胞质外体相关的那些。该酶属于糖基水解酶家族31中的一员，属于α-葡萄糖苷酶系统发育分析的分支1。尽管在体内，金莲花酶催化细胞壁木葡聚糖的动员，但其初级序列与a-葡萄糖苷酶的同源性促使人们研究它对一系列α-葡萄糖苷的作用。它对几种α-(1->；4)和α-(1->；6)连接的底物具有活性，前者的水解速度更快。讨论了这种α-D-木糖苷酶与植物“质外体”α-D-葡萄糖苷酶的功能和进化关系。
We report the isolation, sequencing and analysis of the cDNA corresponding to an a-D-Xylosidase involved in the mobilisation of xyloglucan from the cotyledons of germinated nasturtium (Tropaeolum majus L.) seeds. The translated open reading frame (2,808 bp including the stop codon), gave a polypeptide of 935 amino acids. It included the sequences of eleven peptides obtained by endo-proteinase digestion of the protein, and a putative hydrophobic signal sequence characteristic of a protein that is directed through the plasma membrane. The deduced molecular weight of the translated protein was appreciably higher than the molecular weight determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, suggesting post-translational modification. The protein sequence showed high homology (76.0% identity over 896 amino acids) with a putative a-xylosidase sequence from Arabidopsis thaliana and there was homology with several alpha-glucosidases, notably those associated with the plant cell apoplast. The enzyme is a member of Family 31 of the glycosyl hydrolases and it fits into Clade 1 of the phylogenic analysis of a-glucosidases. Although in vivo the nasturtium enzyme catalyses mobilisation of cell wall xyloglucan, the homology of its primary sequence with a-glucosidases prompted study of its action on a range of alpha-glucosides. It was active against several alpha-(1-->4)-and alpha-(1-->6)-linked substrates, the former being hydrolysed faster. The functional and evolutionary relationships between this a-D-xylosidase and plant "apoplastic" alpha-D-glucosidases are discussed.