Structure of CRISPR associated protein Csn2 from Streptococcus agalactiae

Streptococcus agalactiae, or group B Streptococcus, is the leading cause of bacterial sepsis, pneumonia, and meningitis in newborns in the U.S. and Europe. Beside known cases of antibiotic resistance, Streptococcus also shows resistance against bacteriophages through the recently discovered CRISPR/Cas system. Clustered regularly interspaced short palindromic repeats (CRISPRs) are a family of DNA direct repeats found in many prokaryotic genomes and build up, together with the upstream lying CRISPR-associated (Cas) proteins an adaptive and heritable prokaryotic immune system (1). CRISPR function can be divided into three parts, namely acquisition of new spacers from invading foreign DNA (immunization stage), the expression and processing of CRISPR RNA (crRNA) and CRISPR interference (interference stage). All these steps are directed by Cas proteins, which can be subdivided in 45 gene families associated with CRISPR loci and further subdivided in eight subtypes (2). The Streptococcus agalactiae CRISPR/Cas system belongs to the Nmeni subtype, which is only found within pathogenic bacteria and consists of four different proteins: Csn1, Cas1, Cas2 and Csn2. Csn1 is responsible for CRISPR interference whereas Cas1, Cas2 and Csn2 appear to be involved in generation and integration of new spacers into the CRISPR array by unknown mechanisms (3).

We solved the structure of Csn2 at 2.0Å. The structure shed a light on the function, which could not be deduced from the primary sequence. The monomer of Csn2 contains a helicase domain with a long extended arm, likely used for DNA binding. Furthermore we show that the oligomeric state of the protein is a tetramer and no condition until now was able to abolish this state.