The recombinant dehydrin-like desiccation stress protein from the resurrection plant Craterostigma plantagineum displays no defined three-dimensional structure in its native state
The recombinant dehydrin-like desiccation stress protein from the resurrection plant Craterostigma plantagineum displays no defined three-dimensional structure in its native state
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DOI：
10.1515/bchm3.1996.377.9.555
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发表时间：
1996-09-01
影响因子：
3.7
通讯作者：
Jaenicke, R
中科院分区：
文献类型：
--
作者：
Lisse, T;Bartels, D;Jaenicke, R
耐旱复活植物Craterostigma plantagineum的脱水胁迫伴随着大量脱水胁迫蛋白（Dsp）的积累。其中一个丰富的类别是由Dsp 16蛋白相关的，它含有15个氨基酸的保守的富含赖氨酸的重复序列和一段8个丝氨酸残基，提供了非常亲水的特性。克隆了Craterostigma plantagineum的重组Dsp 16基因，并在大肠杆菌中表达。该蛋白质的纯化和表征其理化性质。无论成功的结晶实验，稀释的水性缓冲溶液不显示一个明确的三维结构的典型的二级结构元素。水溶液中的H-1-NMR（核磁共振）光谱的特征在于未折叠蛋白质典型的小化学位移分散;然而，观察到的线宽对于高度移动的随机大肠杆菌结构不是典型的。相反，它们表明构象状态之间的平衡，优先扩展substructures.As其松散的结构的结果，Dsp 16是非常敏感的蛋白质水解，除非其结构是稳定的结构，使添加剂，如三氟乙醇。变性剂如氯化胍不会诱导协同结构转变。pH依赖性荧光变化反映质子化/去质子化，而不是构象变化。沉降/扩散实验证实了16 kDa的预测分子量。由于高丝氨酸/苏氨酸含量和其松散的结构，Dsp 16易于磷酸化，支持原位结构相对不确定的蛋白质可能参与水结合和磷酸化的想法。
Dehydration stress in the drought-tolerant resurrection plant Craterostigma plantagineum is accompanied by the accumulation of a large number of desiccation stress proteins (Dsp). One abundant class of these is represented by the dehydrin-related Dsp16 protein which contains 15 amino acid conserved lysine-rich repeats and a stretch of eight serine residues providing extremely hydrophilic characteristics. Recombinant Dsp16 from Craterostigma plantagineum has been cloned and expressed in Escherichia coli. The protein was purified and characterized regarding its physicochemical properties.Irrespective of successful crystallization experiments, dilute aqueous buffer solutions do not display a well-defined three-dimensional structure in terms of the canonical secondary structural elements. H-1-NMR (nuclear magnetic resonance) spectra in aqueous solution are characterized by a small chemical shift dispersion typical for an unfolded protein; however, the observed line-widths are not typical for a highly mobile random coli structure. Instead they indicate an equilibrium between conformational states with preferentially extended substructures.As a consequence of its loose structure, Dsp16 is extremely sensitive towards proteolysis unless its structure is stabilized by structure-making additives such as trifluoroethanol. Denaturants such as guanidinium chloride do not induce cooperative structural transitions. pH-dependent fluorescence changes reflect protonation/deprotonation rather than conformational changes. Sedimentation/diffusion experiments confirm the predicted molecular mass of 16 kDa. Due to the high serine/threonine content and its loose structure, Dsp16 is accessible to phosphorylation, supporting the idea that in situ the structurally relatively undefined protein may be involved in both water binding and phosphorylation.