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CRISPR critters: Scientists identify key enzyme in microbial immune system
Date:9/9/2010

A. Cas proteins will use these matching sequences to bind the crRNA to the invading virus or plasmid and silence it."

Haurwitz says the CRISPR/cas system for silencing foreign DNA in prokaryotes is analogous to the way in which short interfering or siRNAs correct genetic problems in eukaryotes. Over time, the CRISPR/cas system will build up inheritable DNA-encoded immunity from future invasions by the same types of viruses and plasmids.

With their crystal structure model of the Csy4 enzyme bound to its cognate RNA, which features a resolution of 1.8 Angstroms, the Berkeley CRISPR research team has shown that Csy4 makes sequence-specific interactions in the major groove of the CRISPR RNA repeat stem-loop. Together with electrostatic contacts to the phosphate backbone, these interactions enable Csy4 to selectively bind to and cleave pre-crRNAs using phylogenetically conserved residues of the amino acids serine and histidine in the active site.

"Our model explains sequence- and structure-specific processing by a large family of CRISPR-specific endoribonucleases," Doudna says.

Doudna and her colleagues produced their 1.8 Angstrom resolution crystallographic structure using the experimental end stations of Beamlines 8.2.1 and 8.3.1 at Berkeley Lab's Advanced Light Source (ALS). Both beamlines are powered by superconducting bending magnets "superbends" and both feature state-of-the-art multiple-wavelength anomalous diffraction (MAD) and macromolecular crystallography (MX) capabilities. Beamline 8.2.1 is part of the suite of protein crystallography beamlines that comprise the Berkeley Center for Structural Biology.

"The ALS and its protein crystallography beamlines continue to be a critical resource for our research, Doudna says.

The crRNAs used by the CRISPR/cas system for the targeted interference of foreign DNA join the growing ranks of small RNA molecules that mediate a variety of processes in both eukaryotes an
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Contact: Lynn Yarris
lcyarris@lbl.gov
510-486-5375
DOE/Lawrence Berkeley National Laboratory
Source:Eurekalert  

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