Structural basis for RNA-duplex recognition and unwinding by the DEAD-box helicase Mssll6p

摘要

Alan Lambowitz及其同事已确定了结合到双链RNA和一个DNA-RNA杂合物上的Mss116(一种酵母DEAD-box蛋白)的结构。DEAD-box蛋白是核酸解旋酶，其功能是展开及重塑RNA和RNA-蛋白复合物。该结构显示这种酶处在一种“展开前”状态，ATP和RNA结合到不同区域:研究人员提出，一个构形变化在展开过程中将它们聚到一起。该结构还显示了这种酶是怎样区分A-形RNA和B-形DNA的。%DEAD-box proteins are the largest family of nucleic acid helicases, and are crucial to RNA metabolism throughout all domains of life. They contain a conserved 'helicase core' of two RecA-like domains (domains (D)l and D2), which uses ATP to catalyse the unwinding of short RNA duplexes by non-processive, local strand separation. This mode of action differs from that of translocating helicases and allows DEAD-box proteins to remodel large RNAs and RNA-protein complexes without globally disrupting RNA structure4. However, the structural basis for this distinctive mode of RNA unwinding remains unclear. Here, structural, biochemical and genetic analyses of the yeast DEAD-box protein Mssl 16p indicate that the helicase core domains have modular functions that enable a novel mechanism for RNA-duplex recognition and unwinding. By investigating D1 and D2 individually and together, we find that Dl acts as an ATP-binding domain and D2 functions as an RNA-duplex recognition domain. D2 contains a nucleic-acid-binding pocket that is formed by conserved DEAD-box protein sequence motifs and accommodates A-form but not B-form duplexes, providing a basis for RNA substrate specificity. Upon a conformational change in which the two core domains join to form a 'dosed state' with an ATPase active site, conserved motifs in Dl promote the unwinding of duplex substrates bound to D2 by excluding one RNA strand and bending the other. Our results provide a comprehensive structural model for how DEAD-box proteins recognize and unwind RNA duplexes. This model explains key features of DEAD-box protein function and affords a new perspective on how the evolutionarily related cores of other RNA and DNA helicases diverged to use different mechanisms.