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“RNA魔剪”新成员“面纱”揭开 定位病变细胞更精准

2019-07-25 05:06:53     来源:中新网     责任编辑:刘欣

  CRISPR-Cas13系统能够靶向编辑RNA,被称为“RNA魔剪”。随着科学家们研究的深入,其家族成员不断壮大。近日,发表在《自然·通讯》的一项成果,揭开了“RNA魔剪”家族最年轻成员“CRISPR-Cas13d”的神秘面纱。

  这项成果来自福建师范大学欧阳松应教授团队。该团队解析了瘤胃球菌的Cas13d蛋白(简称UrCas13d)与crRNA二元复合物高分辨率晶体结构,揭示了UrCas13d加工前体crRNA的酶活性关键位点,鉴定了crRNA的间隔序列和靶向RNA之间的错配耐受性等之前尚未研究清楚的问题。

  “这就像飞机设计工程师获得了设计并制造一架飞机的图纸与工序,为修理或设计出更出色的飞机提供了蓝图。”欧阳松应表示,这为进一步改造Cas13d作为核酸编辑及检测工具,并发挥其在解决人类疾病与健康等问题上的应用潜力,提供了扎实的结构基础和理论依据。  

  “RNA魔剪”应用潜力巨大

  CRISPR-Cas系统是一种原核生物(细菌和古细菌)的免疫防御系统,该系统表达的非编码RNA(crRNA)及其相关Cas蛋白组成的复合物,使得原核生物能够识别并且“撕碎”入侵噬菌体的核酸物质(DNA或者RNA),用以保持自身遗传物质的“纯洁”,在原核生物的进化中起到重要作用。

  “正是由于这种精确地靶向切割特定核酸序列的功能,CRISPR-Cas系统被开发成高效的基因编辑工具,可以对遗传物质DNA或RNA,进行依赖核酸序列信息的精确识别和切割。”欧阳松应说,其中名声在外的CRISPR-Cas9系统可以高效地编辑DNA序列,用于纠正个体细胞内的基因缺陷,给众多遗传疾病和罕见病治疗带来了希望,该系统已经从实验室研究阶段进入到临床研究阶段,具有治愈或预防多种人类疾病的潜力。

  近年来,源于生物学研究及医学领域的巨大应用潜力,一类可以特异地靶向检测与编辑RNA的CRISPR-Cas系统被发现并报道出来,命名为Cas13(包括了a、b、c、d四个家族成员)系统。这一针对RNA的编辑工具可以在RNA水平调节基因的功能,而不会对基因本身造成永久性改变。

  2018年4月,美国索尔克研究所一个课题组和美国Arbor生物技术公司的课题组分别报道了通过生物信息学方法筛选,并经过功能实验验证的新型Cas蛋白,命名为Cas13d。作为最年轻的成员,Cas13d与其他“兄弟姐妹”在功能上有着相似之处,但更有其独特之处。

  “关键在于大小。”欧阳松应介绍,Cas13d的蛋白序列长度约为930个氨基酸,比其他Cas13家族成员明显偏小,比Cas9更是小33%,这种尺寸使其更容易包装到容量较小的载体中,如腺相关病毒(AAV)载体,因此在开发新药、新材料和新的诊断方法上具有广泛应用潜力。

  获得新型“魔剪”高分辨率晶体结构

  结构生物学是分子生物学、生物化学及生物物理学的分支学科,关心的是生物大分子(例如蛋白质分子和核酸分子)的分子三维结构(包括构架和形态),获得它们的结构信息,并研究生物大分子的结构与功能之间的关系。

  “所有的生命活动都是通过各种生物大分子的相互作用来实现,而结构生物学能够解释生物大分子的构象和相互作用的方式,从原子水平揭示生命过程的详细机制,使人们‘知其然而知其所以然’。”欧阳松应告诉记者,目前结构生物学的研究范围已经涉及到大多数重要的生命活动,分子生物学的每一个前沿突破都与结构生物学密切相关。

  在本研究中,欧阳松应课题组将结构生物学与生命科学前沿研究课题结合,解析了UrCas13d蛋白和crRNA二元复合物的高分辨率晶体结构,可以直观看到每一个原子的位置及它们之间的相互作用细节信息。

  “当入侵者到来,CRISPR-Cas系统会在‘指挥官’(前导序列)的调控下转录出两种‘防御材料’,其中一种就是前体crRNA,另一种是我们研究的Cas蛋白。”欧阳松应比喻说。这一研究清楚地解释了Cas13d这把“新型魔剪”,究竟采用了什么样的空间结构布局来加工前体crRNA,并如何结合成熟的crRNA,形成具有识别靶向RNA能力的功能复合物,从而进一步研究清楚该蛋白具有的crRNA依赖的RNase活性的分子机理。

  相关研究表明,Cas13d可做为一种强大RNA编辑工具,用于纠正老年痴呆症患者细胞中引起疾病的蛋白质失衡。同时,Cas13d可以作为一种可操控的RNA结合模块,有效靶向细胞内的RNA转录物,为转录组工程和疾病治疗提供了一个通用平台。

  “该研究揭开了CRISPR-Cas13d系统的神秘面纱,对其技术机制特点有了更深刻理解,有助于这项技术更广泛地应用,从而造福人类。”欧阳松应说。

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