Xu Zhiwei, Escamilla-Treviño Luis, Zeng Lihui, Lalgondar Mallikarjun, Bevan David, Winkel Brenda, Mohamed Ali, Cheng Chi-Lien, Shih Ming-Che, Poulton Jonathan, Esen Asim
Department of Biological Sciences, The University of Iowa, Iowa 52242, USA.
Plant Mol Biol. 2004 May;55(3):343-67. doi: 10.1007/s11103-004-0790-1.
In plants, Glycoside Hydrolase (GH) Family 1 beta -glycosidases are believed to play important roles in many diverse processes including chemical defense against herbivory, lignification, hydrolysis of cell wall-derived oligosaccharides during germination, and control of active phytohormone levels. Completion of the Arabidopsis thaliana genome sequencing project has enabled us, for the first time, to determine the total number of Family 1 members in a higher plant. Reiterative database searches revealed a multigene family of 48 members that includes eight probable pseudogenes. Manual reannotation and analysis of the entire family were undertaken to rectify existing misannotations and identify phylogenetic relationships among family members. Forty-seven members (designated BGLU1 through BGLU47 ) share a common evolutionary origin and were subdivided into approximately 10 subfamilies based on phylogenetic analysis and consideration of intron-exon organizations. The forty-eighth member of this family ( At3g06510; sfr2 ) is a beta -glucosidase-like gene that belongs to a distinct lineage. Information pertaining to expression patterns and potential functions of Arabidopsis GH Family 1 members is presented. To determine the biological function of all family members, we intend to investigate the substrate specificity of each mature hydrolase after its heterologous expression in the Pichia pastoris expression system. To test the validity of this approach, the BGLU44 -encoded hydrolase was expressed in P. pastoris and purified to homogeneity. When tested against a wide range of natural and synthetic substrates, this enzyme showed a preference for beta -mannosides including 1,4- beta -D-mannooligosaccharides, suggesting that it may be involved in A. thaliana in degradation of mannans, galactomannans, or glucogalactomannans. Supporting this notion, BGLU44 shared high sequence identity and similar gene organization with tomato endosperm beta -mannosidase and barley seed beta -glucosidase/ beta -mannosidase BGQ60.
在植物中，糖苷水解酶（GH）家族1的β-糖苷酶被认为在许多不同过程中发挥重要作用，包括对食草动物的化学防御、木质化、种子萌发期间细胞壁衍生寡糖的水解以及活性植物激素水平的控制。拟南芥基因组测序项目的完成使我们首次能够确定高等植物中家族1成员的总数。反复的数据库搜索揭示了一个由48个成员组成的多基因家族，其中包括8个可能的假基因。我们对整个家族进行了人工重新注释和分析，以纠正现有的错误注释并确定家族成员之间的系统发育关系。47个成员（命名为BGLU1至BGLU47）具有共同的进化起源，并根据系统发育分析和内含子-外显子组织的考虑被细分为大约10个亚家族。该家族的第48个成员（At3g06510；sfr2）是一个β-葡萄糖苷酶样基因，属于一个不同的谱系。本文介绍了拟南芥GH家族1成员的表达模式和潜在功能的相关信息。为了确定所有家族成员的生物学功能，我们打算在毕赤酵母表达系统中异源表达每个成熟水解酶后研究其底物特异性。为了测试这种方法的有效性，编码BGLU44的水解酶在毕赤酵母中表达并纯化至同质。当针对多种天然和合成底物进行测试时，该酶表现出对β-甘露糖苷的偏好，包括1,4-β-D-甘露寡糖，这表明它可能参与拟南芥中甘露聚糖、半乳甘露聚糖或葡糖半乳甘露聚糖的降解。支持这一观点的是，BGLU44与番茄胚乳β-甘露糖苷酶和大麦种子β-葡萄糖苷酶/β-甘露糖苷酶BGQ60具有高度的序列同一性和相似的基因组织。
