Program in Systems Biology, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, ON M5G 1X5, Canada; Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada.
Program in Systems Biology, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, ON M5G 1X5, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada.
Mol Cell. 2014 Jan 23;53(2):221-34. doi: 10.1016/j.molcel.2013.12.025.
RNase L is an ankyrin repeat domain-containing dual endoribonuclease-pseudokinase that is activated by unusual 2,'5'-oligoadenylate (2-5A) second messengers and which impedes viral infections in higher vertebrates. Despite its importance in interferon-regulated antiviral innate immunity, relatively little is known about its precise mechanism of action. Here we present a functional characterization of 2.5 Å and 3.25 Å X-ray crystal and small-angle X-ray scattering structures of RNase L bound to a natural 2-5A activator with and without ADP or the nonhydrolysable ATP mimetic AMP-PNP. These studies reveal how recognition of 2-5A through interactions with the ankyrin repeat domain and the pseudokinase domain, together with nucleotide binding, imposes a rigid intertwined dimer configuration that is essential for RNase catalytic and antiviral functions. The involvement of the pseudokinase domain of RNase L in 2-5A sensing, nucleotide binding, dimerization, and ribonuclease functions highlights the evolutionary adaptability of the eukaryotic protein kinase fold.
RNase L 是一种含锚蛋白重复结构域的双末端核糖核酸内切酶-拟激酶，可被不寻常的 2'，5'-寡聚腺苷酸（2-5A）第二信使激活，从而阻止高等脊椎动物中的病毒感染。尽管它在干扰素调节的抗病毒先天免疫中具有重要作用，但人们对其确切的作用机制知之甚少。在这里，我们展示了 RNase L 与天然 2-5A 激活剂结合的 X 射线晶体和小角 X 射线散射结构的功能特征，其中包括有无 ADP 或非水解 ATP 类似物 AMP-PNP 的情况。这些研究揭示了通过与锚蛋白重复结构域和拟激酶结构域的相互作用识别 2-5A，以及核苷酸结合，如何施加刚性交织二聚体构象，这对于 RNase 的催化和抗病毒功能至关重要。RNase L 的拟激酶结构域参与 2-5A 感应、核苷酸结合、二聚化和核糖核酸酶功能，突出了真核蛋白激酶折叠的进化适应性。
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