A Methanococcus maripaludis endoribonuclease cleaves crRNA precursors

The CRISPR/Cas immune systems of Bacteria and Archaea consist of (i) crRNAs, small RNAs containing spacer sequences derived from viruses and conjugative plasmids, and (ii) Cas proteins involved in the acquisition of new spacers, the transcription and processing of the CRISPR locus and the defense against invading mobile genetic elements. One important Cas protein is Cas6, the endoribonuclease that processes crRNA precursors into smaller interfering crRNAs. In Pyrococcus furiosus, Cas6 was shown to cleave CRISPR repeat sequences¹.

Several CRISPR/Cas subtypes exist that differ in Cas protein composition and CRISPR structure. Our computational analyses of the CRISPR/Cas subtype I-B suggests that the open reading frame MmarC5_0767 of Methanococcus maripaludis C5 encodes for a putative crRNA endonuclease albeit having only 12% amino acid identity to P. furiosus Cas6. We established the production and purification of M. maripaludis C5 MmarC5_0767 and could verify that the enzyme displays crRNA precursor cleavage activity. An unusual catalytic triad within the active site of this endonuclease was identified.