The Need for Speed: Run-On Oligomer Filament Formation Provides Maximum Speed with Maximum Sequestration of Activity
The Need for Speed: Run-On Oligomer Filament Formation Provides Maximum Speed with Maximum Sequestration of Activity
复制标题
DOI：
10.1128/jvi.01647-18
复制
发表时间：
2019-03-01
影响因子：
5.4
通讯作者：
Horton, N. C.
中科院分区：
文献类型：
--
作者：
Barahona, Claudia J.;Basantes, L. Emilia;Horton, N. C.
在这里，我们研究了一种在灰色链霉菌中发展起来的不寻常的抗病毒机制。SgrAI是一种II型限制性内切酶，被激活时会形成连续型寡聚体细丝，并具有加速DNA切割活性和扩大DNA序列特异性的双重特性。破坏Run-On寡聚体细丝的突变消除了野生型SgrAI的强大抗噬菌体活性，观察到即使相对温和的破坏也完全取消了这种抗病毒活性，这表明Run-On寡聚体细丝机制赋予的更高速度对其生物学功能至关重要。SgrAI对DNA切割的模拟揭示了这种新描述的酶调节机制相对于更传统机制的动力学优势的来源，以及负责保护宿主基因组免受激活的SgrAI DNA切割活性破坏的隔离效应的来源。SgrAI是一种负责保护宿主细菌免受病毒感染的酶，也是最早利用这种机制的此类酶之一。在本工作中，SgrAI破坏了丝状结构的形成，并测定了其对纯化酶的速度以及它在细胞中的功能的影响。研究发现，即使是减弱但不会破坏细丝形成的微小破坏，也会消除SgrAI保护细胞免受病毒感染的能力，这是其正常的生物学功能。还进行了酶活性的模拟，并显示了与其他已知机制相比，丝状结构如何大大加快了酶的激活速度，以及如何更好地定位其对感兴趣的分子的作用，如侵入噬菌体DNA。
Here, we investigate an unusual antiviral mechanism developed in the bacterium Streptomyces griseus. SgrAI is a type II restriction endonuclease that forms run-on oligomer filaments when activated and possesses both accelerated DNA cleavage activity and expanded DNA sequence specificity. Mutations disrupting the run-on oligomer filament eliminate the robust antiphage activity of wild-type SgrAI, and the observation that even relatively modest disruptions completely abolish this anti-viral activity shows that the greater speed imparted by the run-on oligomer filament mechanism is critical to its biological function. Simulations of DNA cleavage by SgrAI uncover the origins of the kinetic advantage of this newly described mechanism of enzyme regulation over more conventional mechanisms, as well as the origin of the sequestering effect responsible for the protection of the host genome against damaging DNA cleavage activity of activated SgrAI.IMPORTANCE This work is motivated by an interest in understanding the characteristics and advantages of a relatively newly discovered enzyme mechanism involving filament formation. SgrAI is an enzyme responsible for protecting against viral infections in its host bacterium and was one of the first such enzymes shown to utilize such a mechanism. In this work, filament formation by SgrAI is disrupted, and the effects on the speed of the purified enzyme as well as its function in cells are measured. It was found that even small disruptions, which weaken but do not destroy filament formation, eliminate the ability of SgrAI to protect cells from viral infection, its normal biological function. Simulations of enzyme activity were also performed and show how filament formation can greatly speed up an enzyme's activation compared to that of other known mechanisms, as well as to better localize its action to molecules of interest, such as invading phage DNA.