What is CRISPR-Cas9 structure?

Abstract. Many bacterial clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated (Cas) systems employ the dual RNA-guided DNA endonuclease Cas9 to defend against invading phages and conjugative plasmids by introducing site-specific double-stranded breaks in target DNA.

What is the structure of Cas9?

Cas9 features a bi-lobed architecture with the guide RNA nestled between the alpha-helical lobe (blue) and the nuclease lobe (cyan, orange, and gray). These two lobes are connected through a single bridge helix.

Which structure is used to assemble to target specific sequences in the CRISPR-Cas9?

The sgRNA (purple) targets the Cas9 protein to genomic sites containing sequences complementary to the 5′ end of the sgRNA. The target DNA sequence needs to be followed by a proto-spacer adjacent motif (PAM), typically NGG.

What are the three components of CRISPR?

Simplified diagram of a CRISPR locus. The three major components of a CRISPR locus are shown: cas genes, a leader sequence, and a repeat-spacer array.

What is the role of Cas9?

Cas9 is a bacterial RNA-guided endonuclease that uses base pairing to recognize and cleave target DNAs with complementarity to the guide RNA. The programmable sequence specificity of Cas9 has been harnessed for genome editing and gene expression control in many organisms.

What is the function of Cas9 in CRISPR?

When the target DNA is found, Cas9 – one of the enzymes produced by the CRISPR system – binds to the DNA and cuts it, shutting the targeted gene off. Using modified versions of Cas9, researchers can activate gene expression instead of cutting the DNA.

How does the CRISPR-Cas9 process able to be specific for its target?

Scientists are keen to find a way to ensure that the CRISPR-Cas9 binds and cuts accurately. Two ways this may be achieved are through: the design of better, more specific guide RNAs using our knowledge of the DNA sequence of the genome and the ‘off-target’ behaviour of different versions of the Cas9-gRNA complex.

How do the structures of Cas9 help explain the function that each of these proteins plays in the cell?

Cas9 is shaped like a claw. How do the structures of dystrophin and Cas9 help explain the function that each of these proteins plays in the cell? The structure of these proteins is directly related to their function. For dystrophin, it is structured in such a way that it can hold pieces of muscle cells together.

What are the 2 components of CRISPR-Cas9?

In total, the CRISPR-Cas9 system consists of two key components. The first component of the CRISPR-Cas9 system is an RNA molecule known as the guide RNA (gRNA), that can identify the sequence of DNA to be edited. The second component of the CRISPR-Cas9 system is a non-specific CRISPR-associated endonuclease Cas9.

What is the PAM sequence and why is it important for Cas9 activity?

The protospacer adjacent motif (or PAM for short) is a short DNA sequence (usually 2-6 base pairs in length) that follows the DNA region targeted for cleavage by the CRISPR system, such as CRISPR-Cas9. The PAM is required for a Cas nuclease to cut and is generally found 3-4 nucleotides downstream from the cut site.

What is Cas9 and what is its function?