Structural insights into the potential of 4-fluoroproline to modulate biophysical properties of proteins
Structural insights into the potential of 4-fluoroproline to modulate biophysical properties of proteins
复制标题
DOI：
10.1039/c2sc20545a
复制
发表时间：
2012-01-01
期刊：
影响因子：
8.4
通讯作者：
Marx, Andreas
中科院分区：
文献类型：
--
作者：
Holzberger, Bastian;Obeid, Samra;Marx, Andreas
非天然氨基酸4-氟脯氨酸（4-FPro）可用于取代肽和蛋白质中的天然脯氨酸，以改变其稳定性、构象和折叠行为。有趣的是，两种非对映异构体（4 R）-和（4S）-FPro的行为非常不同，导致例如蛋白质稳定性增加或降低。所观察到的相反性质的原因似乎非常复杂，并且尚未得到很好的理解，特别是到目前为止，只有一个4-FPro修饰蛋白质的单一X射线结构可用。这里报道的Taq DNA聚合酶的大片段的晶体结构远远超过了以前研究的蛋白质的分子量和4-FPro残基的数量，并揭示了4-FPro如何影响复杂的蛋白质框架。事实证明，脯氨酰C g -脯氨酰的所有方面都必须综合考虑，以了解它们如何解释蛋白质稳定性的诱导差异。基于新形成的相互作用和4-FPro的构象偏好的不同效应的相互作用决定了可接受的非对映体是否稳定或不稳定靶蛋白。由于平衡效应，4-FPro似乎是一种非常有前途的工具，甚至可以改变具有大量Pro残基的大型酶的性质，因为靶蛋白对多个位点的单核苷酸具有良好的耐受性。值得注意的是，用4-FPro替换Pro显然也导致DNA聚合酶的结晶能力提高。
The unnatural amino acid 4-fluoroproline (4-FPro) can be used to replace natural proline in peptides and proteins to alter their stability, conformation and folding behavior. Interestingly, the two diastereomers (4R)- and (4S)-FPro behave quite differently resulting for example in increased or decreased protein stabilities. The reasons for the observed, opposed properties seem to be very complex and are not well understood yet, especially as only one single X-ray structure of a 4-FPro-modified protein is available, so far. The crystal structure of the large fragment of Taq DNA polymerase reported here far exceeds the molecular mass and number of 4-FPro residues of previous studied proteins and sheds light on how 4-FPro influences complex protein frameworks. It turns out that all aspects of prolyl C g - fluorination have to be considered in a combined fashion to understand how they account for the induced differences in protein stability. The interplay of different effects based on newly formed interactions and on the conformational preferences of 4-FPro determines whether the accepted diastereomer stabilizes or destabilizes the target protein. Due to counterbalanced effects, 4-FPro seems to be a very promising tool to even modify the properties of large enzymes with a high number of Pro residues since mono-fluorination at multiple sites is well tolerated by the target protein. Notably, the replacement of Pro by 4-FPro apparently also led to an improved crystallization capability of the DNA polymerase.