Asset Details
Do you wish to reserve the book?
Structural basis for promiscuous PAM recognition in type I–E Cascade from E. coli
Hey, we have placed the reservation for you!
By the way, why not check out events that you can attend while you pick your title.
Oops! Something went wrong.
Looks like we were not able to place the reservation. Kindly try again later.
Are you sure you want to remove the book from the shelf?
Structural basis for promiscuous PAM recognition in type I–E Cascade from E. coli
Do you wish to request the book?
Structural basis for promiscuous PAM recognition in type I–E Cascade from E. coli
Please be aware that the book you have requested cannot be checked out. If you would like to checkout this book, you can reserve another copy
We have requested the book for you!
Your request is successful and it will be processed during the Library working hours. Please check the status of your request in My Requests.
Oops! Something went wrong.
Looks like we were not able to place your request. Kindly try again later.
Journal Article
Structural basis for promiscuous PAM recognition in type I–E Cascade from E. coli
2016
Overview
The structure of
E. coli
Cascade bound to foreign target DNA is presented, revealing the basis of the relaxed Cascade PAM recognition specificity, which results from its interaction with the minor groove, and demonstrating how a wedge in Cascade forces the directional pairing of the target strand with CRISPR RNA while stabilizing the non-target displaced strand.
Structure of DNA-bound Cascade complex
In the CRISPR system of bacterial immune surveillance, now widely used for genome editing, a CRISPR RNA (crRNA)-bound Cascade complex interacts with double-stranded DNA that can undergo complementary base pairing. The crRNA binds the target strand to form an R-loop structure. The trinucleotide PAM motif near the target sequence is responsible for non-self discrimination. Ailong Ke and colleagues have solved the structure of Cascade bound to foreign target DNA. This reveals the basis of Cascade's relaxed PAM specificity, resulting from its interaction with the minor groove, and shows how a wedge in Cascade forces the directional pairing of the target strand with crRNA, and at the same time stabilizes the non-target, displaced strand.
Clustered regularly interspaced short palindromic repeats (CRISPRs) and the
cas
(CRISPR-associated) operon form an RNA-based adaptive immune system against foreign genetic elements in prokaryotes
1
. Type I accounts for 95% of CRISPR systems, and has been used to control gene expression and cell fate
2
,
3
. During CRISPR RNA (crRNA)-guided interference, Cascade (CRISPR-associated complex for antiviral defence) facilitates the crRNA-guided invasion of double-stranded DNA for complementary base-pairing with the target DNA strand while displacing the non-target strand, forming an R-loop
4
,
5
. Cas3, which has nuclease and helicase activities, is subsequently recruited to degrade two DNA strands
4
,
6
,
7
. A protospacer adjacent motif (PAM) sequence flanking target DNA is crucial for self versus foreign discrimination
4
,
8
,
9
,
10
,
11
,
12
,
13
,
14
,
15
,
16
. Here we present the 2.45 Å crystal structure of
Escherichia coli
Cascade bound to a foreign double-stranded DNA target. The 5′-ATG PAM is recognized in duplex form, from the minor groove side, by three structural features in the Cascade Cse1 subunit. The promiscuity inherent to minor groove DNA recognition rationalizes the observation that a single Cascade complex can respond to several distinct PAM sequences. Optimal PAM recognition coincides with wedge insertion, initiating directional target DNA strand unwinding to allow segmented base-pairing with crRNA. The non-target strand is guided along a parallel path 25 Å apart, and the R-loop structure is further stabilized by locking this strand behind the Cse2 dimer. These observations provide the structural basis for understanding the PAM-dependent directional R-loop formation process
17
,
18
.
Publisher
Nature Publishing Group UK,Nature Publishing Group
Subject
/ Bacterial structural biology
/ Cellular signal transduction
/ Clustered Regularly Interspaced Short Palindromic Repeats - genetics
/ CRISPR-Associated Proteins - chemistry
/ CRISPR-Associated Proteins - metabolism
/ DNA
/ E coli
/ Escherichia coli - chemistry
/ Escherichia coli Proteins - chemistry
/ Escherichia coli Proteins - genetics
/ Escherichia coli Proteins - metabolism
/ Humanities and Social Sciences
/ letter
/ Multiprotein Complexes - chemistry
/ Multiprotein Complexes - metabolism
/ Protein Subunits - chemistry
/ Protein Subunits - metabolism
/ Science
