Information on EC 2.1.1.230 - 23S rRNA (adenosine1067-2'-O)-methyltransferase

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The expected taxonomic range for this enzyme is: Streptomyces

EC NUMBER
COMMENTARY hide
2.1.1.230
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RECOMMENDED NAME
GeneOntology No.
23S rRNA (adenosine1067-2'-O)-methyltransferase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
S-adenosyl-L-methionine + adenosine1067 in 23S rRNA = S-adenosyl-L-homocysteine + 2'-O-methyladenosine1067 in 23S rRNA
show the reaction diagram
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SYSTEMATIC NAME
IUBMB Comments
S-adenosyl-L-methionine:23S rRNA (adenosine1067-2'-O)-methyltransferase
The methylase that is responsible for autoimmunity in the thiostrepton producer Streptomyces azureus, renders ribosomes completely resistant to thiostrepton [2].
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
in the absence of the N-terminal domain, Tsr is catalytically inactive despite possessing the intact S-adenosyl-L-methionine binding sites and catalytic center. The Tsr-C-terminal domain dimer binds the RNA 30fold more weakly than the wild-type enzyme and is unable to promote the N-terminal domain-dependent RNA conformational change
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + adenosine in an RNA fragment containing nucleotides 1029-1122 of the 23S ribosomal RNA from Escherichia coli
S-adenosyl-L-homocysteine + 2'-O-methyladenosine in an RNA fragment containing nucleotides 1029-1122 of the 23S ribosomal RNA from Escherichia coli
show the reaction diagram
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?
S-adenosyl-L-methionine + adenosine1067 in 23S rRNA
S-adenosyl-L-homocysteine + 2'-O-methyladenosine1067 in 23S rRNA
show the reaction diagram
S-adenosyl-L-methionine + adenosine1067 in Escherichia coli 23S rRNA
S-adenosyl-L-homocysteine + 2'-O-methyladenosine1067 in Escherichia coli 23S rRNA
show the reaction diagram
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?
additional information
?
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compared the methylation activity of the Arg135Ala NHR heterodimer with the wild-type NHR homodimer and a series of 29 nt wild-type and mutant RNA substrates. The inactive heterodimer complex binds the RNA more efficiently than the inactive mutant homodimer. Construction of diverse mutant RNA substrates in which A1067 is replaced by adenine structural analogues, A1067G retains significant activity, while 7-deaza-A and 1-methyl-A substitutions at A1067 reduce the activity, overview
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
S-adenosyl-L-methionine + adenosine1067 in 23S rRNA
S-adenosyl-L-homocysteine + 2'-O-methyladenosine1067 in 23S rRNA
show the reaction diagram
COFACTOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
S-adenosyl-L-methionine
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
K+
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ionic optimum: 50-100 mM
NH4+
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ionic optimum: 50-100 mM
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Protein L11
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binding of either thiostrepton or protein L11 inhibits methylation
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S-Adenosyl-D-homocysteine
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competitivewith respect to S-adenosyl-L-methionine
S-adenosyl-L-homocysteine
Thiostrepton
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binding of either thiostrepton or protein L11 inhibits methylation
additional information
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.001
adenosine in an RNA fragment containing nucleotides 1029-1122 of the 23S ribosomal RNA from Escherichia coli
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pH 7.5, 35°C
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0.1 - 0.15
S-adenosyl-L-methionine
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0005 - 0.013
S-adenosyl-L-homocysteine
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
7 - 7.8
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pH 7.0: about 60% of maximal activity, pH 7.8: about 60% of maximal activity
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
30 - 45
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quite active from 30°C-45°C
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
35000 - 38000
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gel filtration
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
SDS-PAGE and gel filtration
homodimer
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structures of the enzyme and the enzyme in complex with S-adenosyl-L-methionine at 2.0 and 2.1 A resolution, respectively
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6
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unstable below
485512
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3)-pLysS by nickel affinity and heparin affinity chromatography, followed by gel filtration
recombinant N-terminally His6-tagged wild-type and mutant enzymes from Escherichia coli by nickel affinity chromatography and gel filtration
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
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gene tsr, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)-pLysS
recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes, with additional FLAG and HA tags, in Escherichia coli
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D36A
mutation significantly decreases activity
E220Q
mutant has 10% of activity compared to wild-type enzyme
E35A
mutation significantly decreases activity
E91A
mutation significantly decreases activity
F88A
mutation significantly decreases activity
H258A/E259A
double mutaion decreases methyltransferase activity to 40% compared to wild-type activity
R165A
mutation significantly decreases activity
R92A
mutation significantly decreases activity
S219A
mutation slightly decreases activity
R162A
site-directed mutagenesis, the mutant binds RNA with 100fold weaker affinity than wild-type Tsr
R26A
site-directed mutagenesis, the mutant is unable to promote the RNase V1-sensitive RNA structural changes, but maintains its interaction with the RNA under certain conditions for the protection of nucleotides G1068 and G1071 from cleavage by RNase T1