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Volume 11
Journal of Proteomics & Bioinformatics
ISSN: 0974-276X
Structural Biology 2018
September 24-26, 2018
September 24-26, 2018 | Berlin, Germany
14
th
International Conference on
Structural Biology
Characterization of new CRISPR/Cas9 system from uncharacterized bacterium
Trung Thanh Thach
1
, Nam Hyeong Kim
1
, Junho Hur
2
and
Yong Ho Kim
1
1
SAINT - Sungkyunkwan University, South Korea
2
Kyung Hee University, South Korea
A
major limitation to expand RNA-guided CRISPR/Cas9 toolkit in genome editing is a sequence-specific recognition
of the protospacer adjacent motif (PAM) at the target DNA site by the Cas9 protein. Exploration and characterization
of new Cas9 ortholog binding distinct PAM sequence would beneficially expand the biological applications of the toolkit.
Here, we identified new CRISPR/Cas9 system from uncharacterized bacterial strain. Analyses of the cleaved double-strand
DNA (dsDNA) using deep sequencing showed that our system recognizes and cleaves target dsDNA with the distinct PAM
sequence, yet previously described. Furthermore, structural analyses using small-angle x-ray scattering combined with binding
affinity assays using bio-layer interferometry and PAM-based target DNA cleavage demonstrated that Arg-1083 and Arg-1116
residues in the Cas9-PAM interacting domain are key determinants of the PAM recognition. Collectively, our finding provides
a novel CRIPRS/Cas9 system together with molecular basic understanding into a RNA-guided the distinct PAM-based target
DNA cleavage, thus expand the target space for CRISPR/Cas9 toolkit applications.
Biography
Trung Thanh Thach received his PhD in Structural Biology from Sungkyunkwan University, Korea in 2015. Thereafter, he was appointed as a Research Professor in
Biotechnology Department, Korea University. Since 2017, he has been employed in Pioneering Nano-based Convergence HRD center, Sungkyunkwan University.
His present research interests include Cas9 protein engineering; structural and functional characterization of new CRISPR-Cas systems.
Trung Thanh Thach et al., J Proteomics Bioinform 2018, Volume 11
DOI: 10.4172/0974-276X-C2-116