|Gene:||casE||Accession Numbers: G7426 (EcoCyc), b2756, ECK2751|
Synonyms: ygcH, cse3, Cas6e
Component of: CRISPR-associated complex for antiviral defense (extended summary available)
CasE is the only Cascade subunit that is necessary and sufficient for cleavage of pre-CRISPR RNA (crRNA). The cleavage reaction is highly specific for the endogenous pre-crRNA [Brouns08].
CasE binds the 3' end of pre-crRNA. CasE connects to the Cascade complex through contacts with CasC (Cas7.1) and CasB (Cas2.1). CasE contains tandem RNA recognition motifs (RRMs) connected by an 8 residue linker. The RRMs form a cleft on one face of the protein that binds a short helix from the first CasC subunit. The opposite face of the protein contacts the pre-crRNA and positions it for cleavage. After cleavage, the mature crRNA remains associated with CasE [Jackson14].
A mutation in the conserved H20 residue of CasE leads to loss of pre-crRNA cleavage [Brouns08].
CasE: "CRISPR-associated" [Brouns08]
Gene Citations: [Pul10]
|Map Position: [2,877,810 <- 2,878,409] (62.03 centisomes)||Length: 600 bp / 199 aa|
Molecular Weight of Polypeptide: 22.293 kD (from nucleotide sequence)
Unification Links: ASAP:ABE-0009042 , DIP:DIP-12124N , EchoBASE:EB2916 , EcoGene:EG13115 , EcoliWiki:b2756 , OU-Microarray:b2756 , PortEco:casE , Pride:Q46897 , Protein Model Portal:Q46897 , RefSeq:NP_417236 , RegulonDB:G7426 , SMR:Q46897 , String:511145.b2756 , UniProt:Q46897
|Biological Process:||GO:0006396 - RNA processing
GO:0051607 - defense response to virus [UniProt-GOA, 2011, Brouns08]
GO:0090305 - nucleic acid phosphodiester bond hydrolysis [UniProt-GOA, 2011]
|Molecular Function:||GO:0003723 - RNA binding
[Jackson14, UniProt-GOA, 2011]
GO:0005515 - protein binding [Jackson14, Babu11]
GO:0016787 - hydrolase activity [UniProt-GOA, 2011, Brouns08]
GO:0004518 - nuclease activity [UniProt-GOA, 2011]
GO:0004519 - endonuclease activity [UniProt-GOA, 2011]
|Cellular Component:||GO:0043234 - protein complex [Jackson14, Babu11]|
|MultiFun Terms:||cell processes → defense/survival|
|Growth Medium||Growth?||T (°C)||O2||pH||Osm/L||Growth Observations|
|LB enriched||Yes||37||Aerobic||6.95||Yes [Gerdes et al., 2003, Comment 1]|
|LB Lennox||Yes||37||Aerobic||7||Yes [Baba et al., 2006, Comment 2]|
|M9 medium with 1% glycerol||Yes||37||Aerobic||7.2||0.35||Yes [Joyce et al., 2006, Comment 3]|
|MOPS medium with 0.4% glucose||Yes||37||Aerobic||7.2||0.22||Yes [Baba et al., 2006, Comment 2]|
Subunit of: CRISPR-associated complex for antiviral defense
Subunit composition of
CRISPR-associated complex for antiviral defense = [CasA][CasB]2[CasC]6[CasD][CasE]
Cascade subunit A = CasA (summary available)
Cascade subunit B = CasB (summary available)
Cascade subunit C = CasC (extended summary available)
Cascade subunit D = CasD (summary available)
crRNA endonuclease = CasE (summary available)
CRISPR (clusters of regularly interspersed short palindromic repeats) loci are associated with defense against foreign DNA and occur widely in bacteria and archaea [Young08]. The CRISPR locus of E. coli K-12 is transcribed into a large pre-crRNA. CasE alone or within the Cascade complex is required for processing of pre-crRNA into the mature crRNA. Processed crRNAs of approximately 57 nt copurify with Cascade. In the presence of Cas3, this complex gives rise to resistance against phages whose genomes have regions of complementarity to elements in the CRISPR repeat [Brouns08].
Cryo-EM structures of the Cascade complex before and after binding to a target sequence have been solved, showing that the bound crRNA is protected from degradation while still being available for base pairing with the target [Wiedenheft11]. Target nucleotide binding results in a concerted conformational change that may result in recruitment of the Cas3 nuclease [Jore11, Wiedenheft11].
The 11 Cas proteins assemble with the crRNA into a 'sea-horse'-shaped structure. CasE forms the head of the complex and binds the 3' end of the pre-crRNA and positions it for cleavage; CasE, CasA and one subunit of CasC form the tail which contacts the 5' end of crRNA. The 6 CasC subuits form a helical backbone which along with the 2 CasB subunits connects the head and tail. The crRNA is interwoven into the Cascade complex in a manner that presents segments for target DNA binding [Jackson14]. The crRNA-target DNA hybrid adopts a ribbon conformation rather than a helix; this conformation is facilitated by the structural disruption of base pairing at every 6th nucleotide in the hybrid molecule. Protein-nucleic acid interactions between the DNA-RNA hybrid and the Cascade complex (in particular the helical backbone) stabilise the ribbon conformation [Mulepati14].
Cas3 as well as a mixture of CasC, CasD and CasE proteins catalyses ATP-independent annealing of RNA with DNA to form hybrid molecules of RNA base-paired into duplex DNA, also known as R-loops [Howard11]. The specificity of the CRISPR system is provided by the formation of R-loops [Jore11]. Strict complementarity between the target and the crRNA is only required in a seven-nucleotide seed region within the protospacer region of crRNAs [Semenova11].
Molecular Weight: 405.0 kD (experimental) [Jore11]
|Biological Process:||GO:0051607 - defense response to virus [Brouns08]|
|Molecular Function:||GO:0003677 - DNA binding
GO:0003723 - RNA binding [Jackson14, Brouns08]
GO:0008270 - zinc ion binding [Jackson14]
GO:0071667 - DNA/RNA hybrid binding [Mulepati14]
|Cellular Component:||GO:0043234 - protein complex [Jackson14]|
Enzymatic reaction of: CRISPR RNA processing (CRISPR-associated complex for antiviral defense)
The reaction direction shown, that is, A + B ↔ C + D versus C + D ↔ A + B, is in accordance with the direction of enzyme catalysis.
The reaction is physiologically favored in the direction shown.
|Chain||2 -> 199|
Markus Krummenacker on Tue Oct 14, 1997:
Gene object created from Blattner lab Genbank (v. M52) entry.
Baba et al., 2006: Baba T, Ara T, Hasegawa M, Takai Y, Okumura Y, Baba M, Datsenko KA, Tomita M, Wanner BL, Mori H (2006). "Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection." Mol Syst Biol 2;2006.0008. PMID: 16738554
Babu11: Babu M, Beloglazova N, Flick R, Graham C, Skarina T, Nocek B, Gagarinova A, Pogoutse O, Brown G, Binkowski A, Phanse S, Joachimiak A, Koonin EV, Savchenko A, Emili A, Greenblatt J, Edwards AM, Yakunin AF (2011). "A dual function of the CRISPR-Cas system in bacterial antivirus immunity and DNA repair." Mol Microbiol 79(2);484-502. PMID: 21219465
Brouns08: Brouns SJ, Jore MM, Lundgren M, Westra ER, Slijkhuis RJ, Snijders AP, Dickman MJ, Makarova KS, Koonin EV, van der Oost J (2008). "Small CRISPR RNAs guide antiviral defense in prokaryotes." Science 321(5891);960-4. PMID: 18703739
Gerdes et al., 2003: Gerdes SY, Scholle MD, Campbell JW, Balazsi G, Ravasz E, Daugherty MD, Somera AL, Kyrpides NC, Anderson I, Gelfand MS, Bhattacharya A, Kapatral V, D'Souza M, Baev MV, Grechkin Y, Mseeh F, Fonstein MY, Overbeek R, Barabasi AL, Oltvai ZN, Osterman AL (2003). "Experimental determination and system level analysis of essential genes in Escherichia coli MG1655." J Bacteriol 185(19);5673-84. PMID: 13129938
Jackson14: Jackson RN, Golden SM, van Erp PB, Carter J, Westra ER, Brouns SJ, van der Oost J, Terwilliger TC, Read RJ, Wiedenheft B (2014). "Crystal structure of the CRISPR RNA-guided surveillance complex from Escherichia coli." Science. PMID: 25103409
Jore11: Jore MM, Lundgren M, van Duijn E, Bultema JB, Westra ER, Waghmare SP, Wiedenheft B, Pul U, Wurm R, Wagner R, Beijer MR, Barendregt A, Zhou K, Snijders AP, Dickman MJ, Doudna JA, Boekema EJ, Heck AJ, van der Oost J, Brouns SJ (2011). "Structural basis for CRISPR RNA-guided DNA recognition by Cascade." Nat Struct Mol Biol 18(5);529-36. PMID: 21460843
Joyce et al., 2006: Joyce AR, Reed JL, White A, Edwards R, Osterman A, Baba T, Mori H, Lesely SA, Palsson BO, Agarwalla S (2006). "Experimental and computational assessment of conditionally essential genes in Escherichia coli." J Bacteriol 188(23);8259-71. PMID: 17012394
PerezRodriguez11: Perez-Rodriguez R, Haitjema C, Huang Q, Nam KH, Bernardis S, Ke A, DeLisa MP (2011). "Envelope stress is a trigger of CRISPR RNA-mediated DNA silencing in Escherichia coli." Mol Microbiol 79(3);584-99. PMID: 21255106
Pul10: Pul U, Wurm R, Arslan Z, Geissen R, Hofmann N, Wagner R (2010). "Identification and characterization of E. coli CRISPR-cas promoters and their silencing by H-NS." Mol Microbiol 75(6):1495-512. PMID: 20132443
Semenova11: Semenova E, Jore MM, Datsenko KA, Semenova A, Westra ER, Wanner B, van der Oost J, Brouns SJ, Severinov K (2011). "Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence." Proc Natl Acad Sci U S A 108(25);10098-103. PMID: 21646539
Wiedenheft11: Wiedenheft B, Lander GC, Zhou K, Jore MM, Brouns SJ, van der Oost J, Doudna JA, Nogales E (2011). "Structures of the RNA-guided surveillance complex from a bacterial immune system." Nature 477(7365);486-9. PMID: 21938068
Arslan13: Arslan Z, Stratmann T, Wurm R, Wagner R, Schnetz K, Pul U (2013). "RcsB-BglJ-mediated activation of Cascade operon does not induce the maturation of CRISPR RNAs in E. coli K12." RNA Biol 10(5). PMID: 23392250
Wade06: Wade JT, Roa DC, Grainger DC, Hurd D, Busby SJ, Struhl K, Nudler E (2006). "Extensive functional overlap between sigma factors in Escherichia coli." Nat Struct Mol Biol 13(9);806-14. PMID: 16892065
Westra10: Westra ER, Pul U, Heidrich N, Jore MM, Lundgren M, Stratmann T, Wurm R, Raine A, Mescher M, Van Heereveld L, Mastop M, Wagner EG, Schnetz K, Van Der Oost J, Wagner R, Brouns SJ (2010). "H-NS-mediated repression of CRISPR-based immunity in Escherichia coli K12 can be relieved by the transcription activator LeuO." Mol Microbiol 77(6);1380-93. PMID: 20659289
Yang14: Yang CD, Chen YH, Huang HY, Huang HD, Tseng CP (2014). "CRP represses the CRISPR/Cas system in Escherichia coli: evidence that endogenous CRISPR spacers impede phage P1 replication." Mol Microbiol 92(5);1072-91. PMID: 24720807
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