Purification crystallization crystallographic analysis and phasing of the CRISPR-associated protein Csm2 from Thermotoga maritima

摘要

The clusters of regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated proteins (Cas) system consists of an intriguing machinery of proteins that confer bacteria and archaea with immunity against phages and plasmids via an RNA-guided interference mechanism. Here, the cloning, recombinant expression in Escherichia coli BL21 (DE3), purification, crystallization and preliminary X-ray diffraction analysis of Csm2 from Thermotoga maritima are reported. Csm2 is thought to be a component of an important protein complex of the type IIIA CRISPR–Cas system, which is involved in the CRISPR–Cas RNA-guided interference pathway. The structure of Csm2 was solved via cadmium single-wavelength anomalous diffraction (Cd-SAD) phasing. Owing to its involvement in the CRISPR–Cas system, the crystal structure of this protein could be of importance in elucidating the mechanism of type IIIA CRISPR–Cas systems in bacteria and archaea.