Membrane Topology and Predicted RNA-Binding Function of the ‘Early Responsive to Dehydration (ERD4)’ Plant Protein

Functional annotation of uncharacterized genes is the main focus of computational methods in the post genomic era. These tools search for similarity between proteins on the premise that those sharing sequence or structural motifs usually perform related functions, and are thus particularly useful for membrane proteins. Early responsive to dehydration (ERD) genes are rapidly induced in response to dehydration stress in a variety of plant species. In the present work we characterized function of Brassica juncea ERD4 gene using computational approaches. The ERD4 protein of unknown function possesses ubiquitous DUF221 domain (residues 312–634) and is conserved in all plant species. We suggest that the protein is localized in chloroplast membrane with at least nine transmembrane helices. We detected a globular domain of 165 amino acid residues (183–347) in plant ERD4 proteins and expect this to be posited inside the chloroplast. The structural-functional annotation of the globular domain was arrived at using fold recognition methods, which suggested in its sequence presence of two tandem RNA-recognition motif (RRM) domains each folded into βαββαβ topology. The structure based sequence alignment with the known RNA-binding proteins revealed conservation of two non-canonical ribonucleoprotein sub-motifs in both the putative RNA-recognition domains of the ERD4 protein. The function of highly conserved ERD4 protein may thus be associated with its RNA-binding ability during the stress response. This is the first functional annotation of ERD4 family of proteins that can be useful in designing experiments to unravel crucial aspects of stress tolerance mechanism.

后基因组时代，计算方法的核心研究方向之一是对功能未表征基因进行功能注释。此类工具基于“共享序列或结构基序的蛋白质通常执行相关功能”这一前提，在蛋白质间搜索同源相似性，因此对膜蛋白的研究尤为实用。早期脱水响应（Early Responsive to Dehydration, ERD）基因可在多种植物物种中被脱水胁迫快速诱导表达。本研究通过计算生物学方法，对芥菜型油菜（Brassica juncea）ERD4基因的功能进行了表征。该功能未知的ERD4蛋白含有普遍存在的DUF221结构域（氨基酸残基312–634位），且在所有植物物种中均保守存在。我们推测该蛋白定位于叶绿体膜中，且至少含有9个跨膜螺旋结构。我们在植物ERD4蛋白中检测到一段由165个氨基酸残基组成的球状结构域（183–347位），并推测该结构域定位于叶绿体基质内。本研究通过折叠识别方法对该球状结构域进行了结构-功能注释，结果显示其序列中含有两个串联的RNA识别基序（RNA-recognition motif, RRM）结构域，每个结构域均折叠为βαββαβ拓扑结构。通过与已知RNA结合蛋白进行基于结构的序列比对，我们发现ERD4蛋白的两个推定RNA识别结构域中，均保守存在两个非经典的核糖核蛋白亚基基序。因此，高度保守的ERD4蛋白的功能，可能与其在胁迫响应过程中的RNA结合能力相关。本研究是首个针对ERD4蛋白家族的功能注释成果，可为设计实验以解析植物胁迫耐受机制的关键环节提供理论支撑。