STRUCTURAL IDENTIFICATION AND CHARACTERIZATION OF PROTEINS INVOLVED IN CRISPR-MEDIATED ANTIVIRAL DEFENCE

Abstract

CRISPR-mediated defense system is known to target and degrade the invading nucleic acids through subsequent infection. In the Type I-E CRISPR-Cas system, Cse2 is proposed to provide a platform to facilitate the targeting of the invading dsDNA by crRNA. Here we report the crystal structure of Meiothermus ruber Cse2 at 2.8 ?. M. ruber Cse2 adopts an a-helical bundle scaffold and harbors a positive surface for nucleic acid binding. Stable M. ruber Cascade is readily formed by co-expression of M. ruber Cascade proteins together with G-rich crRNA in vitro. In addition, we studied the crystal structure of Thermobifida fusca Cse1, which reveals a well-defined dsDNA binding site. The conserved positive patch on the electrostatic surfaces of Cse1 is contiguous with that of Cse2 and adjacent to Cas3, suggesting that Cse1 might be involved in R-loop formation and/or stabilization, as well as the subsequent recruitment of Cas3 for target cleavage.