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Enzyme Nomenclature
Information on EC 2.7.7.52 - RNA uridylyltransferase
for references in articles please use BRENDA:EC2.7.7.52
Word Map on EC 2.7.7.52
- 2.7.7.52
-
uridylylation
- brucei
-
trypanosomes
-
grnas
- kinetoplastids
-
u-insertion
- tarentolae
-
nucleotidyl
- zcchc6
-
u-specific
-
tap-tagged
-
lin28b
-
template-independent
-
precleaved
-
processome
- uridyltransferase
-
maxicircle
-
u-deletion
-
u-tailed
-
pre-let-7
-
dis3l2
- medicine
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The enzyme appears in viruses and cellular organisms
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EC NUMBER 
COMMENTARY 
2.7.7.52
-
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RECOMMENDED NAME
GeneOntology No.
RNA uridylyltransferase
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REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
UTP + RNAn = diphosphate + RNAn+1
enzyme requires an oligoribonucleotide or polyribonucleotide with a free terminal 3'-OH as a primer
-
-
-
UTP + RNAn = diphosphate + RNAn+1
mechanism, uridine insertion in the editing process
-
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
nucleotidyl group transfer
-
-
-
-
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SYSTEMATIC NAME
IUBMB Comments
UTP:RNA uridylyltransferase
The enzyme requires an oligoribonucleotide or polyribonucleotide with a free terminal 3'-OH as a primer.
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CAS REGISTRY NUMBER
COMMENTARY
78519-53-6
-
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ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
editosome protein TbMP57, distinct from mitochondrial enzyme form, i.e. TbTUTase 108
-
-
RET1 and RET2, i.e. RNA editing TUTases 1 and 2, as components of different editing complexes
-
-
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GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
physiological function
-
isoforms TUT7/ZCCHC6, TUT4/ZCCHC11, and TUT2/PAPD4/GLD2 are the terminal uridylyltransferases responsible for pre-miRNA mono-uridylation. The terminal uridylyl transferases act specifically on dsRNAs with a 1 nucleotide 3' overhang, thereby creating a 2 nucleotide 3' overhang. Depletion of terminal uridylyl transferases reduces let-7 microRNA levels and disrupts let-7 function. Although the let-7 suppressor, Lin28, induces inhibitory oligo-uridylation in embryonic stem cells, mono-uridylation occurs in somatic cells lacking Lin28 to promote let-7 biogenesis; isoforms TUT7/ZCCHC6, TUT4/ZCCHC11, and TUT2/PAPD4/GLD2 are the terminal uridylyltransferases responsible for pre-miRNA mono-uridylation. The terminal uridylyl transferases act specifically on dsRNAs with a 1 nucleotide 3' overhang, thereby creating a 2 nucleotide 3' overhang. Depletion of terminal uridylyl transferases reduces let-7 microRNA levels and disrupts let-7 function. Although the let-7 suppressor, Lin28, induces inhibitory oligo-uridylation in embryonic stem cells, mono-uridylation occurs in somatic cells lacking Lin28 to promote let-7 biogenesis
physiological function
-
isoform RET2 is an integral component of the RNA editing core complex RECC. Interaction of RET2 with RECC is accomplished via a protein-protein contact between its middle domain and structural subunit MP81. The recombinant RET2 catalyzes a faithful editing on gapped precleaved double-stranded RNA substrates, and this reaction requires an internal monophosphate group at the 5' end of the mRNA 3' cleavage fragment. RET2 processivity is limited to insertion of three U residues. Incorporation into the RECC voids the internal phosphate requirement and allows filling of longer gaps similar to those observed in vivo. Monomeric and RECC-embedded enzymes display a similar bimodal activity, the distributive insertion of a single uracil is followed by a processive extension limited by the number of guiding nucleotides
physiological function
-
isoform RET1 adds U tails to gRNAs, rRNAs, and selected mRNAs and contributes U residues into A/U heteropolymers. Isoform RET1's terminal uridylyl transferase activity is required for the nucleolytic processing of gRNA, rRNA, and mRNA precursors. The U tails presence does not affect the stability of gRNAs and rRNAs, while transcript-specific uridylylation triggers 3' to 5' mRNA decay. The minicircle-encoded antisense transcripts, which are stabilized by RET1-catalyzed uridylylation, may direct a nucleolytic cleavage of multicistronic precursors
physiological function
isoform Rsp1 associates with uridylyltransferase Rdn1-RNA-dependent RNA polymerase Rdf1 or Rdn1-Rdf2 subcomplexes, creating Rsp1 complexes RSPCs that are physically separate from RNA-dependent RNA polymerase complexes RDRCs. The uridylyltransferase activity of Rdn1 is greatly reduced in RSPCs compared with RDRCs, suggesting enzyme regulation by the alternative partners. Despite the loss of all known RDRC-generated classes of endogenous sRNAs, RSP1 gene knockout is tolerated in growing cells. A minority class of dicer protein Dcr2-dependent sRNAs persists in cells lacking Rsp1 with increased size heterogeneity
physiological function
-
isoforms Zcchc11 and Zcchc6 redundantly control let-7 biogenesis in embryonic stem cells; isoforms Zcchc11 and Zcchc6 redundantly control let-7 biogenesis in embryonic stem cells
physiological function
isoform Tailor preferentially uridylates mirtron hairpins, thereby impeding the production of non-canonical microRNAs. Mirtron selectivity is explained by primary sequence specificity of Tailor, selecting substrates ending with a 3'-guanosine. In contrast to mirtrons, conserved Drosophila precursor micro-RNAs are significantly depleted in 3'-guanosine, thereby escaping regulatory uridylation. Cytoplasmic Tailor is required for miRNA uridylation and normal fertility in flies
physiological function
-
oligo(A)-tailed mRNAs are uridylated by the cytosolic UTP:RNA uridylyltransferase URT1, and URT1 has no major impact on mRNA degradation rates. In absence of uridylation, oligo(A) tails are trimmed, indicating that uridylation protects oligoadenylated mRNAs from 3'-ribonucleolytic attacks. URT1 mutants display an increase in 3'-truncated transcripts
physiological function
isoform HESO1, which uridylates most unmethylated miRNAs in vivo, and isoform URT1 which exhibits nucleotidyl transferase activity on unmethylated miRNA, prefer substrates with different 3'-end nucleotides in vitro and act cooperatively to tail different forms of the same miRNAs in vivo. Both HESO1 and URT1 exhibit nucleotidyl transferase activity on AGO1-bound miRNAs. Although the enzymes are able to add long tails to AGO1-bound miRNAs, the tailed miRNAs remain associated with AGO1. Tailing of AGO1-bound miRNA165/6 drastically reduces the slicing activity of AGO1-miR165/6; URT1 is the single most predominant nucleotidyl transferase that tails miRNAs. URT1 and isoform HESO1, which uridylates most unmethylated miRNAs in vivo, prefer substrates with different 3'-end nucleotides in vitro and act cooperatively to tail different forms of the same miRNAs in vivo. Both HESO1 and URT1 exhibit nucleotidyl transferase activity on AGO1-bound miRNAs. Although the enzymes are able to add long tails to AGO1-bound miRNAs, the tailed miRNAs remain associated with AGO1. Tailing of AGO1-bound miRNA165/6 drastically reduces the slicing activity of AGO1-miR165/6. Monouridylation of miR171a by URT1 endows the miRNA the ability to trigger the biogenesis of secondary siRNAs
physiological function
-
oligo(A)-tailed mRNAs are uridylated by the cytosolic UTP:RNA uridylyltransferase URT1, and URT1 has no major impact on mRNA degradation rates. In absence of uridylation, oligo(A) tails are trimmed, indicating that uridylation protects oligoadenylated mRNAs from 3'-ribonucleolytic attacks. URT1 mutants display an increase in 3'-truncated transcripts
-
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
ATP + 5'(ppp)-UGAGGUAGUAGGUUGUAUAGUU-3'
diphosphate + 5'(ppp)-UGAGGUAGUAGGUUGUAUAGUUA-3'
-
i.e. triphosphorylated human let-7a-5p-3p
-
-
?
ATP + 5'-UGAGGUAGUAGGUUGUAUAGUU-3'
diphosphate + 5'-UGAGGUAGUAGGUUGUAUAGUUA-3'
-
i.e. unphosphorylated human let-7a-0P. Gld2 displays an 83fold preference of ATP over UTP
-
-
?
CTP + 5'(pp)-UGAGGUAGUAGGUUGUAUAGUU-3'
diphosphate + 5'(pp)-UGAGGUAGUAGGUUGUAUAGUUC-3'
-
i.e. diphosphorylated human let-7a-5p-2p
-
-
?
UTP + 5'(p)-UGAGGUAGUAGGUUGUAUAGUU-3'
diphosphate + 5'(p)-UGAGGUAGUAGGUUGUAUAGUUU-3'
-
i.e. monophosphorylated human let-7a-5p. Gld2 displays an 83fold preference of ATP over UTP
-
-
?
UTP + RNAn containing a terminal G residue
diphosphate + RNAn+1
-
substrate dsRNA
-
-
?
ATP + RNAn
diphosphate + RNAn+1
-
editosomal enzyme form, no activity
-
-
-
CTP + RNAn
diphosphate + RNAn+1
-
editosomal enzyme form, no activity
-
-
-
GTP + RNAn
diphosphate + RNAn+1
-
about 50% of the activity with UTP, prefers Mn2+ as divalent cation
-
-
?
GTP + RNAn
diphosphate + RNAn+1
-
editosomal enzyme form, no activity
-
-
-
UTP + RNAn
diphosphate + RNAn+1
-
small guide RNA, i.e. gRNA
-
?
UTP + RNAn
diphosphate + RNAn+1
-
marked specificity for UTP
-
?
UTP + RNAn
diphosphate + RNAn+1
-
responsible for post-transcriptional RNA editing process of mitochondrial transcripts in kinetoplastid protozoans
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
enzyme preferentially uridylates mirtron hairpins
-
?
UTP + RNAn
diphosphate + RNAn+1
-
specifically modifies the 3'-UMP terminal of mammalian U6 small nuclear RNA, i.e. snRNA, structural requirements and specificity, overview
-
?
UTP + RNAn
diphosphate + RNAn+1
-
acts as a host factor to initiate RNA synthesis by poliovirus RNA polymerase in vitro
-
?
UTP + RNAn
diphosphate + RNAn+1
-
RNA substrate specificity, overview
-
?
UTP + RNAn
diphosphate + RNAn+1
-
editosomal enzyme form shows preference for a 3' terminal A or G, while the mitochondrial enzyme form does not
-
?
UTP + RNAn
diphosphate + RNAn+1
-
mitochondrial enzyme form adds Us at the 3' and the 5' end of the RNA
-
?
UTP + RNAn
diphosphate + RNAn+1
-
the mitochondrial enzyme adds a single U to the 3'-end of single-stranded RNA
-
?
UTP + RNAn
diphosphate + RNAn+1
-
involved in uridine insertion in the editing process of RNA
-
?
UTP + RNAn
diphosphate + RNAn+1
-
responsible for post-transcriptional RNA editing process of mitochondrial transcripts in kinetoplastid protozoans
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of primarily single U to single-stranded RNA, addition of the number of Us specified by a guide RNA to insertion editing-like substrates
-
?
UTP + RNAn
diphosphate + RNAn+1
-
TbMP57 TUTase, an enzyme thought to act exclusively in U-insertion, can also function within the U-deletion cycle, after cleavage and 3'-U-exo but before U-deletional cycle
-
-
?
UTP + RNAn
diphosphate + RNAn+1
a template-independent RNA nucleotidyltransferases that specifically recognize UTP, it possesses conserved catalytic and UTP recognition domains. A single nucleoside triphosphate is bound in the active site by a complex network of interactions between amino acid residues, a magnesium ion and highly ordered water molecules with the UTPs base, ribose and phosphate moieties, structure-function analysis, overview
-
-
?
UTP + RNAn
diphosphate + RNAn+1
-
invariant arginine residues 144 and 435 positioned in the vicinity of the UTP-binding site are critical for isoform RET2 activity on single-stranded and double-stranded RNAs, as well as function in vivo. Recognition of a double-stranded RNA, which resembles a guide RNA/mRNA duplex, is further facilitated by multipoint contacts across the RET2-specific middle domain
-
-
?
UTP + RNAn
diphosphate + RNAn+1
-
TbMP57 TUTase, an enzyme thought to act exclusively in U-insertion, can also function within the U-deletion cycle, after cleavage and 3'-U-exo but before U-deletional cycle
-
-
?
UTP + RNAn
diphosphate + RNAn+1
-
the enzyme requires a single-stranded oligoribonucleotide or polyribonucleotide with a free terminal 3'-OH as primer, e.g. oligoA20, tRNAAsp, E. coli RNA, alfalafa mosaic virus RNA 4
-
?
UTP + RNAn
diphosphate + RNAn+1
-
RNA substrate specificity, overview
-
?
UTP + RNAn
diphosphate + RNAn+1
-
RNA uridylyltransferase might function in uridylating specific proteins, RNA is not a natural substrate
-
?
UTP + RNAn containing a terminal U residue
diphosphate + RNAn+1
-
substrate dsRNA
-
-
?
additional information
?
-
enzyme shows strong preference for uridine and a distributive activity for the first added nucleotides. URT1 uridylates oligoadenylated mRNAs
-
-
-
additional information
?
-
HESO1 exhibits nucleotidyl transferase activity on methylated miRNA in vitro. HESO1 has a clear preference for miR158A-U. miR158A-G is the second most preferred substrate
-
-
-
additional information
?
-
URT1 exhibits nucleotidyl transferase activity on unmethylated miRNA in vitro. URT1 shows a strong preference for miR158 ending in A, miR158A-C, miR158A-G, and miR158A-U are similarly used by URT1
-
-
-
additional information
?
-
enzyme shows strong preference for uridine and a distributive activity for the first added nucleotides. URT1 uridylates oligoadenylated mRNAs
-
-
-
additional information
?
-
-
human Gld2 is a bona fide adenylyltransferase with only weak activity toward other nucleotides. Gld2 is a promiscuous enzyme, with activity toward miRNA, pre-miRNA, and polyadenylated RNA substrates. Gld2 shows a clear preference for ATP in the presence of all four NTPs. Apo-Gld2 activity is restricted to adding single nucleotides and processivity likely relies on additional RNA-binding proteins
-
-
-
additional information
?
-
-
enzyme interacts with a minor fraction of total RNA ligase
-
-
-
additional information
?
-
-
post-transcriptional uridylylation of guide RNAs by RNA editing TUTase 1 or RET1, a multi-functional RNA processing enzyme, and U-insertion mRNA editing by RNA editing TUTase 2 or RET2, biological functions of TUT isozymes, RNA processing in mitochondria of trypanosomes, detailed overview
-
-
-
additional information
?
-
-
UTP recognition mechanism, overview. The NTD-CTD bi-domain catalytic modules shared by TUTases and non-canonical poly(A) polymerases, ncPAPs, are quite promiscuous in NTP binding, NTP selectivity of TUTase-like catalytic modules, RNA specificity, overview
-
-
-
additional information
?
-
-
reduction of isozyme RET1 leads to decrease in edited RNA and inhibited growth, lowers the uridine insertion and leads predominantly to shorter gRNAs
-
-
-
additional information
?
-
-
down-regulation of isozyme RET2 inhibits growth and in vivo uridine insertion, but has no effect on the length of gRNAs
-
-
-
additional information
?
-
-
many editing changes are developmentally regulated
-
-
-
additional information
?
-
-
down-regulation of RET1, but not of RET2, affects length distribution of gRNA 3' oligo(U) tails
-
-
-
additional information
?
-
-
Inhibition of RNA editing by down-regulation of expression of the mitochondrial RNA editing TUTase 1 by RNA interference has profound effects on kinetoplast biogenesis in Trypanosoma brucei procyclic cells. De novo synthesis of the apocytochrome b and cytochrome oxidase subunit I proteins is no longer detectable after 3 days of RNAi. The effect on protein synthesis correlates with a decline in the levels of the assembled mitochondrial respiratory complexes III and IV, and also with cyanide-sensitive oxygen uptake. The steady-state levels of nuclear-encoded subunits of complexes III and IV are also significantly decreased. Because the levels of the corresponding mRNAs are not affected, the observed effect is likely due to an increased turnover of these imported mitochondrial proteins. This induced protein degradation is selective for components of complexes III and IV, because little effect is observed on components of the F1-F0 -ATPase complex and on several other mitochondrial proteins
-
-
-
additional information
?
-
-
post-transcriptional uridylylation of guide RNAs by RNA editing TUTase 1 or RET1, a multi-functional RNA processing enzyme, and U-insertion mRNA editing by RNA editing TUTase 2 or RET2, biological functions of TUT isozymes, RNA processing in mitochondria of trypanosomes, detailed overview
-
-
-
additional information
?
-
TUT4 substrate specificity toward nucleoside triphosphate substrates with a synthetic 5'-radiolabeled 24-mer RNA as a primer, in presence of Mg2+ ions TUT4 preferentially incorporates uridylyl residues but the reaction with CTP is also detectable, TbTUT4 incorporates only one deoxynucleotide into RNA, overview. UTP binding site structure and structure-activity analysis
-
-
-
additional information
?
-
-
UTP recognition mechanism, overview. UTP specificity is determined primarily by the two closely positioned carboxylic residues, D297/D421 and E300/E424, which coordinate a crucial water molecule indicated in TUT4/RET2, uracil base interactions with invariant amino acids N147, S148, Y189, D297, E300 of TUT4. The NTD-CTD bi-domain catalytic modules shared by TUTases and non-canonical poly(A) polymerases, ncPAPs, are quite promiscuous in NTP binding, NTP selectivity of TUTase-like catalytic modules, RNA specificity, overview
-
-
-
additional information
?
-
-
isoform RET1 adds U tails to gRNAs, rRNAs, and selected mRNAs and contributes U residues into A/U heteropolymers. Isoform RET1's terminal uridylyl transferase activity is required for the nucleolytic processing of gRNA, rRNA, and mRNA precursors. The U tails presence does not affect the stability of gRNAs and rRNAs, while transcript-specific uridylylation triggers 3' to 5' mRNA decay. The minicircle-encoded antisense transcripts, which are stabilized by RET1-catalyzed uridylylation, may direct a nucleolytic cleavage of multicistronic precursors
-
-
-
additional information
?
-
-
recombinant RET2 catalyzes a faithful editing on gapped precleaved double-stranded RNA substrates, and this reaction requires an internal monophosphate group at the 5' end of the mRNA 3' cleavage fragment. RET2 processivity is limited to insertion of three U residues. Incorporation into the RNA editing core complex RECC allows filling of longer gaps similar to those observed in vivo. Monomeric and RECC-embedded enzymes display a similar bimodal activity, the distributive insertion of a single uracil is followed by a processive extension limited by the number of guiding nucleotides. The distributive +1 insertion creates a substrate for the processive gap-filling reaction. Upon base-pairing of the +1 extended 5' cleavage fragment with a guiding nucleotide, this substrate is recognized by RET2 in a different mode compared to the product of the initial nucleolytic cleavage. Therefore, RET2 distinguishes base pairs in gapped RNA substrates
-
-
-
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NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
UTP + RNAn
diphosphate + RNAn+1
-
responsible for post-transcriptional RNA editing process of mitochondrial transcripts in kinetoplastid protozoans
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
acts as a host factor to initiate RNA synthesis by poliovirus RNA polymerase in vitro
-
?
UTP + RNAn
diphosphate + RNAn+1
-
involved in uridine insertion in the editing process of RNA
-
?
UTP + RNAn
diphosphate + RNAn+1
-
responsible for post-transcriptional RNA editing process of mitochondrial transcripts in kinetoplastid protozoans
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of non-coded poly(U) tail to gRNAs
-
?
UTP + RNAn
diphosphate + RNAn+1
-
addition of primarily single U to single-stranded RNA, addition of the number of Us specified by a guide RNA to insertion editing-like substrates
-
?
UTP + RNAn
diphosphate + RNAn+1
-
RNA uridylyltransferase might function in uridylating specific proteins, RNA is not a natural substrate
-
?
additional information
?
-
-
enzyme interacts with a minor fraction of total RNA ligase
-
-
-
additional information
?
-
-
post-transcriptional uridylylation of guide RNAs by RNA editing TUTase 1 or RET1, a multi-functional RNA processing enzyme, and U-insertion mRNA editing by RNA editing TUTase 2 or RET2, biological functions of TUT isozymes, RNA processing in mitochondria of trypanosomes, detailed overview
-
-
-
additional information
?
-
-
reduction of isozyme RET1 leads to decrease in edited RNA and inhibited growth, lowers the uridine insertion and leads predominantly to shorter gRNAs
-
-
-
additional information
?
-
-
down-regulation of isozyme RET2 inhibits growth and in vivo uridine insertion, but has no effect on the length of gRNAs
-
-
-
additional information
?
-
-
many editing changes are developmentally regulated
-
-
-
additional information
?
-
-
down-regulation of RET1, but not of RET2, affects length distribution of gRNA 3' oligo(U) tails
-
-
-
additional information
?
-
-
Inhibition of RNA editing by down-regulation of expression of the mitochondrial RNA editing TUTase 1 by RNA interference has profound effects on kinetoplast biogenesis in Trypanosoma brucei procyclic cells. De novo synthesis of the apocytochrome b and cytochrome oxidase subunit I proteins is no longer detectable after 3 days of RNAi. The effect on protein synthesis correlates with a decline in the levels of the assembled mitochondrial respiratory complexes III and IV, and also with cyanide-sensitive oxygen uptake. The steady-state levels of nuclear-encoded subunits of complexes III and IV are also significantly decreased. Because the levels of the corresponding mRNAs are not affected, the observed effect is likely due to an increased turnover of these imported mitochondrial proteins. This induced protein degradation is selective for components of complexes III and IV, because little effect is observed on components of the F1-F0 -ATPase complex and on several other mitochondrial proteins
-
-
-
additional information
?
-
-
post-transcriptional uridylylation of guide RNAs by RNA editing TUTase 1 or RET1, a multi-functional RNA processing enzyme, and U-insertion mRNA editing by RNA editing TUTase 2 or RET2, biological functions of TUT isozymes, RNA processing in mitochondria of trypanosomes, detailed overview
-
-
-
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METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mg2+
Mn2+
Mg2+
increases the UTP substrate specificity of TUT4 and influences the enzyme conformation; maximal activity at 2 mM Mg2+
Mg2+
-
divalent cation required, Mg2+ or Mn2+, 4-12 mM stimulate, optimal at 5 mM
Mn2+
-
requires Mn2+ or Mg2+; selectively inhibits the distributive activity of the enzyme; when GTP, CTP, or ATP are used as substrates, Mn2+ is more effective than Mg2+
Mn2+
replacement of magnesium with manganese leads to a dramatic stimulation of processivity and partial loss of selectivity for UTP
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INHIBITORS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Ionic strength
-
quite sensitive to ionic strength, activity decreases by 50% at about 40 mM (NH4)2SO4 or 125 mM potassium acetate
-
additional information
-
RNA interference, i.e. RNAi, induced by tetracyclines causes down-regulation of RET1 and RET2 expression in vitro and in vivo
-
additional information
-
not inhibitory: actinomycin C, rifamycin, alpha-amanitin, phosphate
-
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ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
protein TbMP81
-
editosome protein, interaction enhances the enzyme activity
-
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KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
additional information
steady-state enzyme kinetics, overview
-
0.0004
RNAn
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
0.0025
RNAn
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
0.0126
RNAn containing a terminal U residue
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
-
0.0208
RNAn containing a terminal U residue
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
-
0.0005
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0006
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0025
UTP
-
wild-type, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0026
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0028
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.003
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0081
UTP
-
mutant K149A, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0126
UTP
-
wild-type, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0209
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0003
RNAn
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
0.0007
RNAn
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
0.0015
RNAn containing a terminal U residue
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
-
0.0033
RNAn containing a terminal U residue
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
-
0.00002
UTP
-
mutant K149A, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.00003
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.00005
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.00012
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0003
UTP
-
wild-type, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0012
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0015
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0018
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.003
UTP
-
wild-type, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
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kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.133
RNAn
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
0.167
RNAn
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
0.067
RNAn containing a terminal U residue
-
RNA editing core complex RECC, pH 8.0, temperature not specified in the publication
-
0.267
RNAn containing a terminal U residue
-
recombinant enzyme, pH 8.0, temperature not specified in the publication
-
0.0053
UTP
-
mutant K149A, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0058
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0097
UTP
-
mutant V271R, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.0567
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.0867
UTP
-
mutant C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.117
UTP
-
wild-type, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.21
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal G residue, pH 8.0, temperature not specified in the publication
0.25
UTP
-
wild-type, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
0.6
UTP
-
mutant V271R/C83F, cosubstrate RNAn containing a terminal U residue, pH 8.0, temperature not specified in the publication
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.9
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TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
27
30
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SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
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LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
additional information
additional information
-
integral editosome protein, distinct from mitochondrial enzyme form
-
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PDB
SCOP
CATH
ORGANISM
UNIPROT
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
Trypanosoma brucei brucei (strain 927/4 GUTat10.1)
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MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
500000
700000
-
RNA-stabilized higher complexed form, native PAGE and glycerol gradient sedimentation
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SUBUNITS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
-
a single C2H2-type zinc finger domain of Zcchc11 is responsible for the functional interaction with pluripotency factor Lin28. Lin28 uses terminal uridylyltransferases Zcchc11 and Zcchc6 to control let-7 expression and has important implications for stem cell biology as well as cancer
additional information
isoform Rsp1 associates with uridylyltransferase Rdn1-RNA-dependent RNA polymerase Rdf1 or Rdn1-Rdf2 subcomplexes, creating Rsp1 complexes RSPCs that are physically separate from RNA-dependent RNA polymerase complexes RDRCs. The uridylyltransferase activity of Rdn1 is greatly reduced in RSPCs compared with RDRCs, suggesting enzyme regulation by the alternative partners
additional information
TbTUT4 represents a minimal catalytically active RNA uridylyltransferase consisting of only two domains that define the catalytic center at the bottom of the nucleoside triphosphate and RNA substrate binding cleft. The enzyme shows two significantly different conformations: one molecule is in a relatively open apo conformation, whereas the other displays a more compact TUTase-UTP complex, structure-function analysis, overview
additional information
-
in RET1, an extra 150 amino acid insertion constitutes a helix-rich domain essential for activity, the insertion is designated as middle domain, occurs at a conserved site within the catalytic domain and is found in RET1, RET2 and TUT3, but not in TUT4, analysis of Structural diversity of trypanosomal TUTases, overview
additional information
-
isoform RET2 is an integral component of the RNA editing core complex RECC
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Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purified recombinant N-terminally His-tagged TUT4, free or with bound 2'-deoxyribonucleoside, at a concentration of 10 mg/mL in 10 mM HEPES, pH 7.6, 70 mM KCl, 0.5 mM DTT, and in presence of 4 mM MgCl2 and 0.05 mM UTP, screening with commercial crystallization screens, vapor diffusion technique, 4°C, cryoprotection by 15% glycerol, X-ray diffraction structure determination and analysis at 2.0-2.4 A resolution; small plate-like co-crystals of purified recombinant N-terminally His-tagged TbTUT4 and UTP grow in 100 mM sodium cacodylate (pH 6.5), 200 mM calcium acetate, 18% (w/v) PEG-8000, crystal strucuture reveals two significantly different conformations of this TUTase:one molecule is in a relatively open apo confirmation, whereas the other displays a more compact TUTase-UTP complex
structure of apoenzyme and in complex with UTP, to 1.56 A and 1.95 A resolution, respectively. Enzyme possesses a bridging domain, which extends from the C-terminal domain and makes hydrophobic contacts with the N-terminal domain, thereby creating a cavity adjacent to the UTP-binding site. Enzyme shows no appreciable conformational change upon UTP binding and apparently does not require RNA substrate to select a cognate nucleoside triphosphate.
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TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
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GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
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STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
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Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
expression and purification of 20S editosome-associated RET2 complex from Trypanosoma brucei by affinity isolation; recombinant RET1 from Trypanosoma brucei is purified by metal affinity chromatography (Talon metal affinity column), anion-exchange chromatography (HiTrap Q column) and loading onto a MonoS 5/5 column (final purification step), expected yield is 0.5 mg/liter of bacterial culture, it is important to maintain the concentration of KCl above 80 mM during purification, as protein tends to precipitate under low salt conditions; the recombinant expressed RET2 is purified by loading the protein extract onto a Talon affinity column and onto a HiTrap Sepharose S column, the expected protein yield is 1 mg of protein per liter of bacterial culture
-
homogenous RET1 is purified from an enriched mitochondrial fraction of Leishmania tarentolae by a series of chromatographic steps: affinity chromatography (polyU Sepharose 4B), anion-exchange chromatography (Poros 20 HQ 46/100 column), gel filtration (Superose 12 and Superose 6 columns) and hydrophobic interactions chromatography (Phenyl Superose column); recombinant RET1 from Leishmania tarentolae is purified by cation-exchange resin (Sepharose S Fast Flow), metal affinity chromatography (Talon metal affinity column), anion-exchange chromatography (Mono Q 5/5 column), expected yield: 0.2-0.5 mg/liter
-
partial purification of U6 snRNA specific enzyme form via chromatography on a U6 snRNA affinity resin, separation from unspecific enzyme form
-
recombinant His-tagged wild-type and mutant TUT4 from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, ion exchange chromatography, and gel filtration
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Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
DNA sequence determination, editosomal enzyme, expression as His-tagged protein in Escherichia coli
-
DNA sequence determination, gene is located on chromosome 9, expression as His-tagged protein in Escherichia coli
-
DNA sequence determination, mitochondrial enzyme, expression as His-tagged protein in Escherichia coli
-
expressed in Escherichia coli BL-21 as a 6xHis tagged fusion protein; TUT4, DNA and amino acid sequence determination and analysis, expression of N-terminally His-tagged wild-type and mutant TUT4 in Escherichia coli strain BL21(DE3)
expression in Escherichia coli BL21(DE3) Codon Plus RIL, use of strains bearing additional tRNAs for expression of genes with G-rich codons is essential for RET1 expression
-
expression in Escherichia coli BL21(DE3) Codon Plus RIL, use of strains bearing additional tRNAs for expression of genes with G-rich codons is essential for RET1 expression; for recombinant expression in Escherichia coli BL21 the RET2 gene lacking 21 codons at the 5' end, which correspond to the mitochondria importation signal, is cloned into pET15+vector to generate N-terminal 6 His-RET2 fusion protein; TbRET2 fused with the affinity TAP tag at the C-terminus using pLew79 vector in 29-13 cell line
-
expression of RET2 in Leishmania tarentolae cells as cytosolic enzyme by removing the mitochondrial targeting signal sequence
-
phylogenetic analysis, sequence comparison and signature motifs, overview
phylogenetic analysis, sequence comparison and signature motifs, overview
-
phylogenetic analysis, sequence comparison and signature motifs, overview
-
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ENGINEERING
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D491A/D493A
R121A
Km=0.0028 (UTP), kcat=0.000033/sec (UTP),Km=0.132 (RNA), kcat= 0.005/sec (RNA)
R141A
R307A
Km=0.0004 (UTP), kcat=0.000133/sec (UTP), Km=0.01 (RNA), kcat= 0.00833/sec (RNA)
S148A
Km=0.0873 (UTP), kcat=0.021/sec (UTP), Km=0.0002 (RNA), kcat= 0.0166/sec (RNA)
S188A
Km=0.034 (UTP), kcat=0.0005/sec (UTP), Km=0.0003 (RNA), kcat= 0.00166/sec (RNA)
V271R/C83F
-
240% of wild-type activity for transfer to terminal U residue
additional information
-
mutations of metal coordinating catalytic aspartic residues D66, D68 and D136, render TUT4 inactive but have no effect on UTP binding
R141A
Km=0.001 (UTP), kcat=0.00011/sec (UTP),Km=0.0726 (RNA), kcat= 0.0216/sec (RNA)
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
medicine
-
a single C2H2-type zinc finger domain of Zcchc11 is responsible for the functional interaction with pluripotency factor Lin28. Lin28 uses terminal uridylyltransferases Zcchc11 and Zcchc6 to control let-7 expression and has important implications for stem cell biology as well as cancer; pluripotency factor Lin28 uses terminal uridylyltransferases Zcchc11 and Zcchc6 to control let-7 expression and has important implications for stem cell biology as well as cancer
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LINKS TO OTHER DATABASES (specific for EC-Number 2.7.7.52)
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