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

Author(s)

Gallo, Gloria ; Augusto, Gilles ; Rangel, Giulliana ; Zelanis, Andre ; Mori, Marcelo A. ; Campos, Claudia Barbosa ; Wuertele, Martin

• Document type: Journal article
• Source: ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS; v. 71, p. 5-pg., 2015-10-01.

Abstract

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. (AU)