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Publication Abstract from PubMed

The RNA-guided DNA endonuclease Cas9 cleaves double-stranded DNA targets complementary to an RNA guide, and is widely used as a powerful genome-editing tool. Here, we report the crystal structure of Brevibacillus laterosporus Cas9 (BlCas9, also known as BlatCas9), in complex with a guide RNA and its target DNA at 2.4-A resolution. The structure reveals that the BlCas9 guide RNA adopts an unexpected architecture containing a triple-helix, which is specifically recognized by BlCas9, and that BlCas9 recognizes a unique N(4)CNDN protospacer adjacent motif through base-specific interactions on both the target and non-target DNA strands. Based on the structure, we rationally engineered a BlCas9 variant that exhibits enhanced genome- and base-editing activities with an expanded target scope in human cells. This approach may further improve the performance of the enhanced BlCas9 variant to generate useful genome-editing tools that require only a single C PAM nucleotide and can be packaged into a single AAV vector for in vivo gene therapy.

Structure and engineering of Brevibacillus laterosporus Cas9.,Nakane T, Nakagawa R, Ishiguro S, Okazaki S, Mori H, Shuto Y, Yamashita K, Yachie N, Nishimasu H, Nureki O Commun Biol. 2024 Jul 3;7(1):803. doi: 10.1038/s42003-024-06422-z. PMID:38961195^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.