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SpCas9 major
domains, colored by domain:
Blue
for RuvC
Light
gray for Argenine helix (Arg), equivalent to FnCas9 BH
Dark
gray for Helical domains, equivalent to FnCas9 REC1-3
Green
for HNH
Black
for C-Terminal Domain (CTD), equivalent to FnCas9 WED/PI
SpCas9 has two
"lobes." Arg and Helical domains comprise the REC lobe, while
RuvC, HNH and CTD comprise the NUC (nuclease) lobe.
RuvC
active site (slab view) for cleavage of non-target DNA strand.
Aspartate 10 (orange), glutamate 762 (green), and aspartate 986
(red) coordinate with an Mg2+ ion (not crystallized), while
histidine 983 catalyzes hydrolysis of the DNA backbone. In this
conformation, the active site is too far from the DNA for
hydrolysis.
HNH
active site for cleavage of target DNA strand. Similarly to RuvC,
aspartate 839 (green) and asparagine 863 (orange) bind an Mg2+
ion, while histidine 840 (cyan) performs the DNA cleavage. Like in
RuvC, the HNH active site is in a poor conformation for DNA
hydrolysis, and for Mg2+ binding.
11. F. Jiang, D.W. Taylor, J.S. Chen, J.E.
Kornfeld, K. Zhou, A.J. Thompson, E. Nogales, J.A. Doudna.
“Structures
of a CRISPR-Cas9 R-loop complex primed for DNA cleavage.”
Science 351.6275 (2016): 867-871 (PubMed)
5f9r (PDB)
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