Information on EC 3.1.21.3 - type I site-specific deoxyribonuclease

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY
3.1.21.3
-
RECOMMENDED NAME
GeneOntology No.
type I site-specific deoxyribonuclease
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
endonucleolytic cleavage of DNA to give random double-stranded fragments with terminal 5'-phosphates; ATP is simultaneously hydrolysed
show the reaction diagram
-
-
-
-
endonucleolytic cleavage of DNA to give random double-stranded fragments with terminal 5'-phosphates; ATP is simultaneously hydrolysed
show the reaction diagram
kinetic model for DNA-translocation and DNA cleavage
-
endonucleolytic cleavage of DNA to give random double-stranded fragments with terminal 5'-phosphates; ATP is simultaneously hydrolysed
show the reaction diagram
monitoring of mechanism on 27 MHz quartz-crystal microbalance
-
REACTION TYPE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
hydrolysis of phosphoric ester
-
-
-
-
SYNONYMS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
adenosine triphosphate-dependent deoxyribonuclease
-
-
-
-
ATP-dependent deoxyribonuclease
-
-
-
-
ATP-dependent DNase
-
-
-
-
BsaHI restriction-modification system
B0LX59
-
BsaHI restriction-modification system
Geobacillus stearothermophilus CPW11
B0LX59
-
-
deoxyribonuclease (ATP- and S-adenosyl-L-methionine dependent)
-
-
-
-
deoxyribonuclease (ATP-dependent)
-
-
-
-
DNase
-
-
-
-
EC 3.1.23
-
-
-
-
EC 3.1.24
-
-
-
-
EC 3.1.4.33
-
-
formerly
-
Eco377I
-
-
Eco585I
-
-
Eco646I
-
-
Eco777I
-
-
EcoA0ORF42P
-
-
EcoA0ORF42P
Escherichia coli A0 34/86 (O83:K24:H31)
-
-
-
EcoAI
Q07736
-
EcoAI
Escherichia coli A0 34/86 (O83:K24:H31)
Q07736
-
-
EcoAO83I
Q07736, Q47281
-
EcoAO83I
Escherichia coli A0 34/86 (O83:K24:H31)
Q07736, Q47281
-
-
EcoB
-
-
-
-
EcoEI
Q47281
-
EcoEI
Escherichia coli A0 34/86 (O83:K24:H31)
Q47281
-
-
EcoKI type I DNA restriction enzyme
-
-
EcoprrI
-
-
EcoR124/3I
-
-
EcoR124I
Q304R3
-
EcoR124II
-
-
EcoRII modification enzyme
-
-
EcoRII RM gene complex
-
-
EcoRII system
-
-
Endodeoxyribonuclease
-
-
-
-
exodeoxyribonuclease
-
-
-
-
H91_orf206
-
-
-
-
H91_orf376
-
-
-
-
HpyAXII restriction-modification system
-
-
HpyAXII restriction-modification system
Helicobacter pylori NSH57
-
-
-
KpnBI
Q6WN39
-
MpnORFDAP
-
-
-
-
MpnORFDBP
-
-
-
-
nuclease, deoxyribo-
-
-
-
-
nuclease, deoxyribo-, ATP-dependent
-
-
-
-
PspGI endonuclease
-
-
PspGI restriction-modification system
-
-
R. BsaHI
Geobacillus stearothermophilus CPW11
B0LX59
-
-
R.EcoAI
-
-
-
-
R.EcoEI
-
-
-
-
R.EcoKI
-
-
-
-
R.EcoR124I restriction endonuclease
-
-
-
-
R.EcoR124II
-
-
-
-
R.HpyAXII
-
-
R.HpyAXII
Helicobacter pylori NSH57
-
-
-
R.PspGI
-
-
restriction-modification system
-
-
-
-
Sau1 type I restriction-modification system
-
-
StyLTIII
Salmonella-Escherichia coli hybrid
-
-
StySEAI
Salmonella-Escherichia coli hybrid
-
-
StySGI
Salmonella-Escherichia coli hybrid
-
-
StySKI
Salmonella-Escherichia coli hybrid
-
-
StySPI
Salmonella-Escherichia coli hybrid
-
-
type I R-M enzyme
-
-
type I R-M system
-
-
type I R-M system
Escherichia coli A0 34/86 (O83:K24:H31)
-
-
-
type I restriction enzyme
-
-
-
-
type I restriction enzyme
Escherichia coli, Salmonella-Escherichia coli hybrid
-
-
type I restriction enzyme
-
-
type I restriction enzyme
Q7MPU7
-
type I restriction modification enzyme
-
-
type I restriction-modification enzyme
-
-
type I restriction-modification enzyme
Q304R3
-
type I restriction-modification system
-
-
type I restriction-modification system
Escherichia coli A0 34/86 (O83:K24:H31)
-
-
-
type IB restriction enzyme
Q07736, Q47281
-
type IB restriction enzyme
Escherichia coli A0 34/86 (O83:K24:H31)
Q07736, Q47281
-
-
type II R-M system
-
-
type II R-M system
Helicobacter pylori NSH57
-
-
-
type II restriction-modification system
-
-
KpnBI
Klebsiella pneumoniae GM236
Q6WN39
-
-
additional information
-
a complete listing of all these enzymes and their recognition sites has been produced by R.J. Roberts, this list is updated annually
additional information
-
27 isoschizomers identified among 26 clinical Escherichia coli strains. Without exception, all 26 strains produce one or two type I enzymes. The two most common isoschizomers are Eco377I and Eco646I, which between them account for 48% (13/27) of the R-M systems identified in these strains. The classical type I enzymes (EcoBI, EcoKI and EcoAI) accounted for only 30% (8/27) in clinical samples
additional information
-
a complete listing of all these enzymes and their recognition sites has been produced by R.J. Roberts, this list is updated annually
CAS REGISTRY NUMBER
COMMENTARY
37263-09-5
-
ORGANISM
COMMENTARY
LITERATURE
SEQUENCE CODE
SEQUENCE DB
SOURCE
clinical isolate contains four type I restriction enzymes: Eco394I, Eco826I, Eco851I, Eco912I
-
-
Manually annotated by BRENDA team
EcoR124I
-
-
Manually annotated by BRENDA team
enzyme EcoDXXI
-
-
Manually annotated by BRENDA team
enzyme EcoK; enzyme EcoR
-
-
Manually annotated by BRENDA team
enzyme EcoKI
-
-
Manually annotated by BRENDA team
enzyme EcoR124 and EcoDXXI
-
-
Manually annotated by BRENDA team
enzyme EcoR124I
-
-
Manually annotated by BRENDA team
enzyme isoforms EcoKI, EcoAI, EcoR124I
-
-
Manually annotated by BRENDA team
enzyme R.EcoR124I
-
-
Manually annotated by BRENDA team
enzyme REcoK
-
-
Manually annotated by BRENDA team
enzyme sEcoAI, EcoBI, EcoDI, EcoDXXI, EcoEI, EcoKI, EcoR124I and enzyme EcoR124/3I
-
-
Manually annotated by BRENDA team
enzymes EcoKI, EcoR124I, EcoA1
-
-
Manually annotated by BRENDA team
HsdR subunit of isoform EcoR124I
-
-
Manually annotated by BRENDA team
HsdR subunit of isoform EcoR124I
SwissProt
Manually annotated by BRENDA team
HsdR subunit, type IA restriction-modification complex enzyme EcoKI
-
-
Manually annotated by BRENDA team
recombinant
-
-
Manually annotated by BRENDA team
strain A0 34/86 (O83:K24:H31)
-
-
Manually annotated by BRENDA team
strain A0 34/86 (O83:K24:H31)
UniProt
Manually annotated by BRENDA team
strains 2739, L4001, 1225, EC377 ,EC585, EC646, EC777, EC394 and EC912
-
-
Manually annotated by BRENDA team
Escherichia coli A0 34/86 (O83:K24:H31)
strain A0 34/86 (O83:K24:H31)
-
-
Manually annotated by BRENDA team
Escherichia coli A0 34/86 (O83:K24:H31)
strain A0 34/86 (O83:K24:H31)
UniProt
Manually annotated by BRENDA team
Geobacillus stearothermophilus CPW11
strain CPW11
UniProt
Manually annotated by BRENDA team
strains 26695, J99, HPAG1, NSH57and NSH79
-
-
Manually annotated by BRENDA team
type I restriction-modification enzyme subunit S; GM236
SwissProt
Manually annotated by BRENDA team
Klebsiella pneumoniae GM236
type I restriction-modification enzyme subunit S; GM236
SwissProt
Manually annotated by BRENDA team
several strains isolated from infected potato plant. Presence of enzyme Pca17AI, an isoschizomer of EcoRII endonuclease is only observed in isolates of Pectobacterium carotovorum atrospeticum, not in Pectobacterium carotovorum carotovorum
-
-
Manually annotated by BRENDA team
enzyme s StySBI, StySJI, StySPI and enzyme StySQI
-
-
Manually annotated by BRENDA team
Salmonella-Escherichia coli hybrid
strains L4029 and L4039
-
-
Manually annotated by BRENDA team
strain NCTC8325-4, strain SH1000, strain COL and strain 879R4RF
-
-
Manually annotated by BRENDA team
strain RN4220 can accept plasmid DNA from Escherichia coli. A mutation in hsdR subunit of isoform Sau1 is responsible for this phenotype and its complementation restores a nontransformable phenotype. Complemented strain RN4220 is resistant to bacteriophage lysis if the phage is grown on staphylococcus aureus of a different lineage
-
-
Manually annotated by BRENDA team
strain RN450
-
-
Manually annotated by BRENDA team
Staphylococcus aureus RN450
strain RN450
-
-
Manually annotated by BRENDA team
SUBSTRATE
PRODUCT                      
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
3'-biotinylated 51 bp dsDNA + H2O
?
show the reaction diagram
-
-
-
?
5'-biotinylated 51 bp ssDNA + H2O
?
show the reaction diagram
-
-
-
?
DNA + H2O
?
show the reaction diagram
-
-
-
-
?
DNA + H2O
?
show the reaction diagram
-
-
-
-
?
DNA + H2O
?
show the reaction diagram
B0LX59
-
-
-
?
DNA + H2O
?
show the reaction diagram
Q7MPU7
-
-
-
?
DNA + H2O
?
show the reaction diagram
-
EcoR124I couples ATP hydrolysis to bidirectional DNA translocation
-
-
?
DNA + H2O
?
show the reaction diagram
-
the enzyme must overcome a similar slow step before translocation reaches a steady state
-
-
?
DNA + H2O
?
show the reaction diagram
-
R.HpyAXII effectively restricts chromosomal DNA during natural transformation
-
-
?
DNA + H2O
?
show the reaction diagram
-
HsdR subunit can produce only a single cleavage of the phosphodiester backbone of the DNA but can cooperate with another HsdR subunit to produce full DNA cleavage, producing DNA with overhanging ends of single-stranded DNA. On a single-site plasmid, cleavage requires the association of HsdR, from solution, to the cleavage complex in order to produce double-strand cleavage
-
-
?
DNA + H2O
?
show the reaction diagram
-
the PspGI restriction-modification system recognizes the sequence CCWGG. R.PspGI cuts DNA before the first C in the cognate sequence and M.PspGI methylates N4 of one of the cytosines in the sequence. R.PspGI flips both bases in the central base pair out of the duplex
-
-
?
DNA + H2O
?
show the reaction diagram
Helicobacter pylori NSH57
-
R.HpyAXII effectively restricts chromosomal DNA during natural transformation
-
-
?
DNA + H2O
?
show the reaction diagram
Geobacillus stearothermophilus CPW11
B0LX59
-
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
-
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
-
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
-
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
-
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-, Q07736, Q47281
-
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
overview of recognition sequences
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
overview of recognition sequences
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
the wild type enzyme EcoK cleaves circular DNA. Only one endonuclease molecule is required per cleavage event. Cleavage of linear DNA may require a second endonuclease molecule
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
restricts unmodified phage DNA
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
the enzyme is both a restriction endonuclease and a modification methylase. Hemi-methylated DNA is the preferred substrate for methylation
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
the site of restriction cleavage is random, occuring between 1 and 5 kb from the recognition site. The modification methylase acts directly at the recognition sequence
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
the modification methylase binds sequence specifically to DNA and protects a 25bp fragment containing its cognate recognition sequence from digestion by exonuclease III, specific adenine on each strand of DNA is the site of methylation
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
supercoiled with one or two SR124I recognition sites is cleaved by the same mechanism. Nicked-circle DNA is an intermediate of the cleavage reaction
-
-
-
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
EcoAI preferentially generates 3'-overhangs of 2-3 nt. Displays some preference for the formation of 5'-overhangs of a length of 6-7 and 3-5 nt, respectively. type I restriction enzymes require two restriction subunits to introduce DNA double-stran breaks, each providing one catalytic center for phosphodiester bond hydrolysis, EcoKI displays some preference for the formation of 5'-overhangs of a length of 6-7 and 3-5 nt, respectively. Type I restriction enzymes require two restriction subunits to introduce DNA double-stran breaks, each providing one catalytic center for phosphodiester bond hydrolysis, EcoR124I displays some preference for the formation of 5'-overhangs of a length of 6-7 and 3-5 nt, respectively. Type I restriction enzymes require two restriction subunits to introduce DNA double-stran breaks, each providing one catalytic center for phosphodiester bond hydrolysis
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
initiation of translocation by type I restriction-modification enzymes is associated with a short DNA extrusion
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
-
the two motor subunits of Eco124I are independent motors that translocate along the helical pitch of the DNA. Dynamic termination and reinitiation of translocation activity is governed by disassembly and reassembly of the enzyme
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
Staphylococcus aureus RN450
-
restricts unmodified phage DNA
-
-
-
plasmid DNA + H2O
?
show the reaction diagram
-
R.HpyAXII effectively restricts unmethylated plasmid during natural transformation
-
-
?
plasmid DNA + H2O
?
show the reaction diagram
-
EcoKI prefers to have a partially filled DNA-binding site rather than one fully occupied by non-specific DNA. Dimerization of EcoKI does not occur before DNA binding and takes place on specific sites before any looping. Dimerization occurs before the two specific sites are bought together. Looping initially occurs between a target site and a non-specific region of DNA
-
-
?
plasmid DNA + H2O
?
show the reaction diagram
Helicobacter pylori NSH57
-
R.HpyAXII effectively restricts unmethylated plasmid during natural transformation
-
-
?
synthetic oligonucleotide + ATP
?
show the reaction diagram
-
recognition sequence: CA(underlined)C(5N)T(underlined)GGC, recognition sequence: GA(underlined)C(5N)RT(underlined)AAY, recognition sequence: GCA(underlined)(6N)CT(underlined)GA, recognition sequence: GTCA(underlined)(6N)T(underlined)GAY
-
-
?
duplex DNA + ATP
double-stranded DNA fragments with terminal 5'-phosphate + ADP + inorganic phosphate
show the reaction diagram
Escherichia coli A0 34/86 (O83:K24:H31)
-, Q07736, Q47281
-
-
-
?
additional information
?
-
-
-
-
-
-
additional information
?
-
-
the database REBASE contains information about recognition sites and cleavage sites
-
-
-
additional information
?
-
-
restricts unmodified phage DNA after both infection and transfection
-
-
-
additional information
?
-
-
a bacterial population may switch the recognition sequence of its type I restriction-modification system by single recombination events and thus is able to maintain a prokaryotic analogue of the immune system of variable specificity
-
-
-
additional information
?
-
-
enzyme restricts the exchange of genetic material between bacteria of different strains or species
-
-
-
additional information
?
-
-
isoform Sau1 is the major mechanism for blocking transfer of resistance genes and other mobile genetic elements into Staphylococcus aureus isolates from other species, as well as for controlling the spread of resistance genes between isolates of different Staphylococcus aureus lineages
-
-
-
additional information
?
-
-
basal transcription from promoter esp1396ICR results in gradual accumulation of C.Esp1396I, which upon dimerization binds to a single site in promoter esp1396IM, preventing further transcription from this promoter. Further accumulation of C.Esp1396I results in activation and then gradual repression of promoter esp1396ICR
-
-
-
additional information
?
-
-
C.PvuII binding to operator left activates transcription. Binding preference for operator left over operator right. All four bases of the inter-operator TGTA spacer are required for C.PvuII-dependent activation
-
-
-
additional information
?
-
Staphylococcus aureus RN450
-
restricts unmodified phage DNA after both infection and transfection
-
-
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
(Substrate)
LITERATURE
(Substrate)
COMMENTARY
(Product)
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
plasmid DNA + H2O
?
show the reaction diagram
Helicobacter pylori, Helicobacter pylori NSH57
-
R.HpyAXII effectively restricts unmethylated plasmid during natural transformation
-
-
?
DNA + H2O
?
show the reaction diagram
Helicobacter pylori, Helicobacter pylori NSH57
-
R.HpyAXII effectively restricts chromosomal DNA during natural transformation
-
-
?
additional information
?
-
-
-
-
-
-
additional information
?
-
-
restricts unmodified phage DNA after both infection and transfection
-
-
-
additional information
?
-
-
a bacterial population may switch the recognition sequence of its type I restriction-modification system by single recombination events and thus is able to maintain a prokaryotic analogue of the immune system of variable specificity
-
-
-
additional information
?
-
-
enzyme restricts the exchange of genetic material between bacteria of different strains or species
-
-
-
additional information
?
-
-
isoform Sau1 is the major mechanism for blocking transfer of resistance genes and other mobile genetic elements into Staphylococcus aureus isolates from other species, as well as for controlling the spread of resistance genes between isolates of different Staphylococcus aureus lineages
-
-
-
additional information
?
-
-
basal transcription from promoter esp1396ICR results in gradual accumulation of C.Esp1396I, which upon dimerization binds to a single site in promoter esp1396IM, preventing further transcription from this promoter. Further accumulation of C.Esp1396I results in activation and then gradual repression of promoter esp1396ICR
-
-
-
additional information
?
-
-
C.PvuII binding to operator left activates transcription. Binding preference for operator left over operator right. All four bases of the inter-operator TGTA spacer are required for C.PvuII-dependent activation
-
-
-
additional information
?
-
Staphylococcus aureus RN450
-
restricts unmodified phage DNA after both infection and transfection
-
-
-
COFACTOR
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
ATP
-
is degraded during reaction; required
ATP
-
is degraded during reaction; required
ATP
-
; required
S-adenosyl-L-methionine
-
activity is stimulated by S-adenosyl-L-methionine
METALS and IONS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
Mg2+
-
required
INHIBITORS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
ArdA
-
antirestriction protein of InvB plasmid R16, selective
-
S-adenosyl homocysteine
-
competitive
DNA
-
cleavage of DNA is inhibited by an increased degree of negative supercoiling
additional information
-
a repressing complex of C.Esp1396I tetramer bound to both binding sites
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
S-adenosyl methionine
-
stimulates DNA cleavage activity
S-adenosyl-L-methionine
-
stimulates, not required
S-adenosyl-L-methionine
-
required
S-adenosyl-L-methionine
-
-
S-adenosyl-L-methionine
-
required
additional information
-
REcBCD displaces EcoR124I from the cleaved DNA, allowing it to regain catalytic function and cleave the second aliquot of DNA. RecBCD is able to reactivate cleavage-inactivated EcoR124I holoenzyme
-
additional information
-
an active R.HpyAXII endonuclease can only exist in the presence of its active DNA MTase partner
-
KM VALUE [mM]
KM VALUE [mM] Maximum
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
0.000015
-
3'-biotinylated 51 bp dsDNA
-
pH 9.4, 30C
-
0.000011
-
5'-biotinylated 51 bp ssDNA
-
pH 9.4, 30C
-
TURNOVER NUMBER [1/s]
TURNOVER NUMBER MAXIMUM[1/s]
SUBSTRATE
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
IMAGE
167
-
ATP
-
at 37C
pH OPTIMUM
pH MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
6
-
-
EcoB, methylase activity
8
-
-
nuclease reaction
TEMPERATURE OPTIMUM
TEMPERATURE OPTIMUM MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
80
-
-
optimal temperature for DNA cleavage. Optimal flipping occurs at temperatures substantially below the growth temperature of the source organism for PspGI and for the catalytic activity of endonuclease
LOCALIZATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
individual subunits HsdR and HsdM are soluble cytoplasmic proteins
Manually annotated by BRENDA team
-
associated to, intact enzyme complex, R subunits of EcoKI and EcoR124I are partly accessible at external surface of cytoplasmic membrane, EcoAI subunits are not accessible
Manually annotated by BRENDA team
-
the EcoR124I holoenzyme is particularly exposed on the periplasmic side of the cytoplasmic membrane
Manually annotated by BRENDA team
MOLECULAR WEIGHT
MOLECULAR WEIGHT MAXIMUM
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
17920
-
-
sedimentation equilibrium analytical ultracentrifugation of C.Esp1396I
18450
-
-
theoretical Mr of C.Esp1396I
290000
315000
-
pentameric enzyme form R2M2S1, gel filtration
312000
-
-
gel filtration
400000
-
-
EcoK, sucrose density gradient sedimentation
SUBUNITS
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
?
-
x * 116000, HsdR, x * 55000, HsdM, x * 43000, HsdS
?
-
x * 60000, HsdM subunit, crystallography
?
Q7MPU7
x * 90000, full length protein, SDS-PAGE. X * 60000, crystallized HsdR subunit, SDS-PAGE. X * 95000, putative HsdR subunit, calculated from sequence
dimer
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sedimentation equilibrium analytical ultracentrifugation of C.Esp1396I
octamer
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alpha2,beta4,gamma2, 2 * 135000 + 4 * 60000 + 2 * 55000, EcoB, SDS-PAGE
pentamer
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alpha2,beta2,gamma1, 2 * 135000 + 2 * 62000 + 1 * 52000, EcoK, SDS-PAGE
pentamer
Q304R3
crystallography
monomer
Q304R3
1 * 94000, dynamic light scattering, 1 * 130000, analytical ultracentrifugation, subunit HsdR, subunit is globular and fairly compact
additional information
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the stoichiometry of R.EcoR124 appears to be R1M2S1, the stoichiometry of R.EcoKI is R2M2S1. S is the HsdS-subunit which is responsible for DNA recognition, R is HsdR-subunit which is required for restriction and M is an independent methyltransferase, Mtase
additional information
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ATPase activity, DNA translocase activity and endonuclease activity are specified by the R subunit, the R subunit comprises several different functional domains
additional information
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two enzyme species, the larger species has the stoichiometry R2M2S1, the smaller species has the stoichiometry of R1M2S1. Only the R2M2S1 complex is capable of DNA cleavage, the R1M2S1 complex retains ATPase activity. The HsdS subunit determines specificity, the HsdM subunit is responsible for DNA methylation, the HsdR subunit is required for restriction. The HsdM subunit and the HsdS subunit can also form an independent DNA methyltransferase with a subunit stoichiometry of M2S1
additional information
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the HsdR subunit may be limiting within the cell. An excess of HsdM and HsdS may produce the methylase in vivo and the assembly of the endonuclease may be dependent upon the prior production of this methylase
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
phosphoprotein
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HsdR subunit of isoform EcoKI is phosphorylated on Thr. Phosphorylation in vitro is strictly dependent on the addition of a catalytic amount of cytoplasmic fraction isolated from Escherichia coli. Phosphorylation in vivo only occurs when subunit HsdR is coproduced with subunits HsdM and HsdS
Crystallization/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
database information: http://rebase.neb.com
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model of subunit HsdR using protein fold-recognition and homology modeling. Subunit shows an ellipsoidal shape of the enzymatic core comprising the N-terminal and central domains. Conformational heterogenity of the C-terminal region implicated in binding of HsdR to the HsdS-HsdM complex
Q304R3
motor subunit HsdR of pR124 plasmid-borne type I restriction-modification enzyme EcoR124I, solved in complex with Mg2+-ATP at 2.6 A resolution. HsdR presents four globular domains in a square-planar arrangement, generating prominent grooves between adjacent domain pairs. Lys220 on alpha8 is 3.1 A from N3 on the exposed edge of ATP bound at the helicase domains, potentially coupling endonuclease and helicase function. A uniformly positive surface groove with a clear match to the size and shape of duplex DNA proceeds from a canonical helicase cleft in a continuous path down the front of the motor subunit between the helical and endonuclease domains, where the cleavage site is recessed slightly from the surface
Q304R3
recombinant motor subunit HsdR of isoform EcoR124I in presence and absence of ATP, at 2.6 A resolution
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hanging-drop and sitting-drop vapour-diffusion method, HsdR subunit crystallized from 8%(w/v) polyethylene glycol 3350, 0.15 M ammonium chloride, 0.1 M HEPES pH 7.5 and 2 mM beta-mercaptoethanol, to 2.60 A resolution. Crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 71.01, b = 89.04, c = 113.66 A. With one HsdR molecule in the asymmetric unit, the Matthews coefficient is 2.14 A3 Da-1 and the solvent content is 42%
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HsdM subunit, to 1.86 A resolution, by hanging-drop and sitting-drop vapour-diffusion methods. Crystal belongs to the tetragonal space group P41212 or P43212, with unit-cell parameters a = b = 78.9, c = 165.8 A. With one molecule in the asymmetric unit, the crystal volume per unit protein weight is 2.12 A3/Da, with a solvent content of 42%
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N-terminal fragment (ca. 590 amino acids) of a putative HsdR subunit (817 amino acids), native crystals and Se-Met crystals, to 2.5 A resolution, by hanging-drop vapor-diffusion method at 22C. Se-Met crystal belongs to the orthorhombic space group P212121 with unit-cell dimensions of a = 71.01, b = 89.04 and c = 113.66 A. Seven Se sites in the asymmetric unit at 3.0 A resolution. Crystal structure reveals catalytic sites for nuclease (NTD-domain, Ala21-Ile159) and ATPase (RecA-like domains: RD1 for Lys160-Tyr360 and RD2 for Ala372-Val590) activities and suggests a possible translocation mechanism of the HsdR subunit
Q7MPU7
Purification/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
enzyme EcoB; enzyme EcoK
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mixture of two enzyme species, the larger species has the stoichiometry R2M2S1, the smaller species has the stoichiometry of R1M2S1.only the R2M2S1 complex is capable of DNA cleavage, the R1M2S1 complex retains ATPase activity
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Mtase(DELTA50)
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mutant methylase
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recombinant motor subunit HsdR of isoform EcoR124I
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wild-type R.PspGI and its mutant purified by chromatography, to homogeneity
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C.Esp1396I-6His and C.Esp1396I without hexahistidine tag
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HsdM subunit, on anion-exchange column, more than 95% pure
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HsdR fusion protein purified by gel filtration. HsdR with five additional amino acids (YFQGA) at the N-terminus purified on anion-exchange column and by gel filtration, purified protein is more than 95% pure
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native HsdR protein homogeneously purified by sequential chromatographic steps
Q7MPU7
Cloned/COMMENTARY
ORGANISM
UNIPROT ACCESSION NO.
LITERATURE
248-bp insertion fragments of the ptypeI plasmid inserted at the HincII site of pUC19
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; recombinant plasmid pJS4M overproducing HsdM compared to HsdS
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EcoR124I HsdR with selenomethionine labeling expressed in Escherichia coli
Q304R3
enzyme with and without a temperature sensitive mutation in the hsdS gene are cloned in pBR322 plamid and introduced into Escherichia coli C3-6
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expressed in Escherichia coli strains BNH670 or GM31 harboring a plasmid with various versions of the EcoRII RM gene complex
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HsdR subunit of isoform EcoR124I, expression in Escherichia coli
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of hsdS(DELTA50)
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transforming the Escherichia coli BL21(DE3) strains with a BAC C4/1 carrying the hsdR, hsdM and hsdS genes of EcoAO83I and with plasmids carrying the hsdS and hsdM genes of EcoAI
-, Q07736, Q47281
PCR-amplified gene for C.PvuII cloned downstream of the arabinose-inducible PBAD promoter in vector pBAD24 yielding plasmids pIM1
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His-tagged gene cloned into the NheI/EcoRI sites of the pTXBI vector and expressed in Escherichia coli ER2566
B0LX59
pCR-TOPO/hpyAXII digested with EcoRI and subcloned into plasmid Bluescript SK+. Plasmid pET15b/hpyAXIIR expressed in Escherichia coli strain BL21CodonPlus(DE3)-RIL
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wild-type R.PspGI and its mutant expressed as pET21a-PspGI-WT and pET21a-PspGI-D138A in Escherichia coli strain ER2744
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hsdR PCR product cloned into pMAD, giving rise to pDELTAHsdR-1. Plasmid electroporated into the transformable strain RN4220 at 30C, with erythromycin selection, and subsequently transduced to NCTC8325-4, SH1000 and COL using phage 80alpha
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esp1396I genes expressed in Escherichia coli HB101 and XL1-Blue
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full length HsdR transformed into Escherichia coli B834(DE3) cells
Q7MPU7
HsdR subunit cloned into pProExHTc and expressed in Escherichia coli B834(DE3)
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putative HsdM subunit amplified, cloned into pProExHTc, which expresses 25 extra amino acids containing six consecutive His residues at the N-terminus. The expression construct transformed into Escherichia coli B834 (DE3)
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ENGINEERING
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
T239C
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shows decreased DNA methyltransferase activity at a higher temperature in vivo and in vitro than the nonmutated enzyme
T402C
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cells carrying the mutation are able to grow at 42C
D354A
B0LX59
activity is significantly impaired
E350A
B0LX59
activity is significantly impaired
F353A
B0LX59
activity is significantly impaired
P349A
B0LX59
activity is significantly impaired
Q344A
B0LX59
similar activity to the wild-type
R351A
B0LX59
activity is negligible
R352A
B0LX59
similar activity to the wild-type
S348A
B0LX59
similar activity to the wild-type
E350A
Geobacillus stearothermophilus CPW11
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activity is significantly impaired
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P349A
Geobacillus stearothermophilus CPW11
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activity is significantly impaired
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Q344A
Geobacillus stearothermophilus CPW11
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similar activity to the wild-type
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R352A
Geobacillus stearothermophilus CPW11
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similar activity to the wild-type
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S348A
Geobacillus stearothermophilus CPW11
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similar activity to the wild-type
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L80P
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L80P mutation in the modification enzyme of the EcoRII gene complex confers thermosensitivity of cell growth (shows activity at 30C but not at 37C). Under a condition of inhibited protein synthesis, the activity of the mutant is completely lost at a high temperature. In parallel, the L80P mutant protein disappears more rapidly than the wild-type protein
additional information
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a temperature-sensitive mutation within the hsdS gene appears to affect the ability of the HsdR subunit to interact with the HsdS subunit when forming an active endonuclease
additional information
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mutation in which the specificity has altered due to a Tn5 insertion into the middle of the hsdS, the gene which encodes the polypeptide that confers DNA sequence specificity to both the restriction and the modification reaction. The mutant recognition sequence is an interrupted palindrome, TCA(N8)TGA, in which the 5' half site of the wild type site is repeated in inverse orientation. The additional base pair in the non-specific spacer of the mutant recognition sequence maintains the proper spacing between the two methylatable adenine groups. Tn5 insertion occurs at nucleotide 673 of the 1221 bp gene. This effectively deletes the entire carboxyl-terminal DNA binding domain which recognizes the 3' half of the EcoDXXI binding site. The truncated hsdS gene still encodes both the amino-terminal DNA binding domain and the conserved repeated sequence that defines the length of the recognition site spacer region
additional information
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N-terminal 83-amino-acid deletion mutant, shows activity at at 30C and 37C. The EcoRII RM gene complex is a mutant carrying a T239C (L80P) substitution in the modification gene and a synonymous T402C substitution in the restriction gene. The capability of the restriction-modification system EcoRII in forcing maintenance on its host can be modulated by a region of its antitoxin, the modification enzyme, as in the classical postsegregational killing systems
additional information
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point mutation within the hsdS gene or insertional mutagenesis using the Tn5 transposon, both show new DNA specificities. Both new specificities are due to the production of a truncated HsdS subunit, which is able to dimerize, producing the recognition sequence GAAn7TTC in the case of the point mutation of EcoR124I, which produces a stop codon and hence the truncated subunit
D138A
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catalytically inactive
additional information
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gene of subunit hsdR from strain RN4220 contains a premature stop codon resulting in a truncated R.Sau1 product of 192 amino acids. Strain RN4220 can accept plasmid DNA from Escherichia coli. The truncated hsdR subunit is responsible for this phenotype and its complementation restores a nontransformable phenotype. Complemented strain RN4220 is resisitant to bacteriophage lysis if the phage is grown on staphylococcus aureus of a different lineage
additional information
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hsdR null mutants, complete deletion of the hsdR gene is not sufficient to generate readily transformable NCTC8325-4, SH1000 and COL strains. HsdR mutant strains have growth rates identical to the parental strains and and are competent if transformed with appropriately modified Staphylococcus aureus DNA. HsdR mutants are competent but do not accept foreign DNA due to the presence of additional factor(s) that degrade unmodified DNA. Heat treatment of competent cells increases transformation efficiency of hsdR mutants
additional information
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mutations within the tetranucleotide repeats diminish binding of C.Esp1396I. Insertion of an additional base into the TATA sequence to make the GACT/AGCT sequences symmetric around a TAT spacer reduces DNA binding
APPLICATION
ORGANISM
UNIPROT ACCESSION NO.
COMMENTARY
LITERATURE
biotechnology
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potential uses for EcoR124I as a nanoactuator within a biosensor in the field of bionanotechnology. It may be used as a molecular dynamo that can measure events from single molecules of DNA, providing a highly sensitive biosensor for detecting events that interrupt translocation events
medicine
-, Q07736, Q47281
identification of a putative restriction-modification system EcoA0ORF42P in the commensal Escherichia coli strain A0 34/86 (O83: K24: H31), which is efficiently used at Czech paediatric clinics for prophylaxis and treatment of nosocomial infections and diarrhoea of preterm and newborn infants. This type I R-M system is a member of the type IB family, but is not an isoschizomer of either any prototype of the type IB members or any sequenced putative IB R-M systems. It is designated as EcoAO83I, the DNA recognition sequence of the EcoAO83I is GGA(8N)ATGC. Combination of the classical biochemical and bacterial genetics approaches with comparative genomics may contribute effectively to further classification of many other putative type-I enzymes, especially in clinical samples
medicine
Escherichia coli A0 34/86 (O83:K24:H31)
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identification of a putative restriction-modification system EcoA0ORF42P in the commensal Escherichia coli strain A0 34/86 (O83: K24: H31), which is efficiently used at Czech paediatric clinics for prophylaxis and treatment of nosocomial infections and diarrhoea of preterm and newborn infants. This type I R-M system is a member of the type IB family, but is not an isoschizomer of either any prototype of the type IB members or any sequenced putative IB R-M systems. It is designated as EcoAO83I, the DNA recognition sequence of the EcoAO83I is GGA(8N)ATGC. Combination of the classical biochemical and bacterial genetics approaches with comparative genomics may contribute effectively to further classification of many other putative type-I enzymes, especially in clinical samples
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additional information
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inactivation of the hsdR gene of the Sau1 type I restriction-modification system is responsible for the high transformation efficiency of RN4220
additional information
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transcription of both the restriction endonuclease and methyltransferase genes is controlled by interdependent regulatory loops governed by the C.Esp1396I protein