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Information on EC 2.7.7.79 - tRNAHis guanylyltransferase and Organism(s) Saccharomyces cerevisiae and UniProt Accession P53215
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2.7.7.79 tRNAHis guanylyltransferaseIUBMB Comments
In eukarya an additional guanosine residue is added post-transcriptionally to the 5'-end of tRNAHis molecules. The addition occurs opposite a universally conserved adenosine73 and is thus the result of a non-templated 3'-5' addition reaction. The additional guanosine residue is an important determinant for aminoacylation by EC 6.1.1.21, histidine---tRNA ligase.The enzyme requires a divalent cation for activity . ATP activation is not required when the substrate contains a 5'-triphosphate (ppp-tRNAHis) .
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Word Map
- 2.7.7.79
- guanosine
- polymerases
- polymerization
-
histidyl-trna
- aminoacylation
-
fidelity
-
anticodon
-
post-transcriptionally
-
watson-crick
-
template-dependent
-
nucleotidyl
-
genome-encoded
- methanosarcina
- ntp
- acetivorans
-
phosphodiester
- thuringiensis
- medicine
The taxonomic range for the selected organisms is: Saccharomyces cerevisiae
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
hide(Overall reactions are displayed. Show all >>)
p-tRNAHis
+
+
+
=
pGp-tRNAHis
+
+
2
p-tRNAHis
+
=
App-tRNAHis
+
App-tRNAHis
+
=
pppGp-tRNAHis
+
Synonyms
trna(his) guanylyltransferase, trnahis guanylyltransferase, thg1l, hsthg1, thg1p, mathg1, hthg1, bttlp, histidine trna guanylyltransferase, pathg1, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
p-tRNAHis + ATP + GTP + H2O = pGp-tRNAHis + AMP + 2 diphosphate
overall reaction
SYSTEMATIC NAME
IUBMB Comments
p-tRNAHis:GTP guanylyltransferase (ATP-hydrolysing)
In eukarya an additional guanosine residue is added post-transcriptionally to the 5'-end of tRNAHis molecules. The addition occurs opposite a universally conserved adenosine73 and is thus the result of a non-templated 3'-5' addition reaction. The additional guanosine residue is an important determinant for aminoacylation by EC 6.1.1.21, histidine---tRNA ligase.The enzyme requires a divalent cation for activity [2]. ATP activation is not required when the substrate contains a 5'-triphosphate (ppp-tRNAHis) [3].
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
p-tRNAHis + ATP + dGTP
?
dGTP is almost as efficient as GTP for the guanylylation process. While GDP still serves for the enzymatic reaction, GMP is accepted very poorly by the enzyme
-
-
?
p-tRNAHis + ATP + GDP
?
dGTP is almost as efficient as GTP for the guanylylation process. While GDP still serves for the enzymatic reaction, GMP is accepted very poorly by the enzyme
-
-
?
p-tRNAHis + ATP + GTP
pppG-p-tRNAHis + AMP + diphosphate
the extra nucleotide in position -1 of mitochondrial and eukaryotic cytoplasmic tRNAHis molecules is added posttranscriptionally to the 5' end of the tRNA by a histidine-tRNA specific guanylyltransferase
-
-
?
p-tRNAHis + dATP + GTP
pppGp-tRNAHis + dAMP + diphosphate
30% of the activity compared to ATP
-
-
?
ppp-tRNALeu + GTP
?
the D68A mutation causes a dramatic decrease in the rigorous specificity of Thg1 for tRNAHis. This single alteration enhances the kcat/KM for ppp-tRNALeu by 38-fold relative to that of wild-type Thg1
-
-
?
ppp-tRNAPhe + GTP
?
the D68A mutation causes a dramatic decrease in the rigorous specificity of Thg1 for tRNAHis. This single alteration enhances the kcat/KM for ppp-tRNAPhe by nearly 100-fold relative to that of wild-type Thg1
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
-
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
the G(-1) residue is both necessary and sufficient for aminoacylation of tRNA by histidyl-tRNA synthetase in vitro and is required for aminoacylation in vivo
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
among all acceptor tRNAs in unfractionatedt RNA only tRNAHis is a substrate for the purified enzyme. tRNA from plant and prokaryotes are better substrates than mammalian and insect tRNAs
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
during the first step ATP is cleaved to AMP and diphosphate creating adenylylated enzyme. In a second step the activated enzyme forms a stable complex with its cognate tRNA substrate. The 5'-phosphate of the tRNA is adenylylated by nucleotide transfer from the adenylylated guanylyltransferase to form A(5')pp(5') at the 5'-end of the tRNA. Finally, the 3'-hydroxyl of GTP adds to the activated 5' terminus of the tRNA with the release of AMP. dGTP is almost as efficient as GTP for the guanylylation process. While GDP still serves for the enzymatic reaction, GMP is accepted very poorly by the enzyme
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
specific for tRNAHis. No activity with tRNAPhe. No ATP is required when ppp-tRNAHis is used as a substrate
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
Thg1 is more than 10000-fold more selective for its cognate substrate tRNAHis than for the noncognate substrate tRNAPhe. Alteration of this anticodon in tRNAHis completely abrogates Thg1 activity, and the introduction of this GUG anticodon to tRNAPhe or tRNAGly results in significant Thg1 activity
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
-
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
overall reaction
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
overall reaction
-
-
?
additional information
?
-
in vivo Thg1 catalyzes 3'5' polymerization on tRNAHisC73, but not on tRNAHisA73
-
-
?
additional information
?
-
-
in vivo Thg1 catalyzes 3'5' polymerization on tRNAHisC73, but not on tRNAHisA73
-
-
?
additional information
?
-
retention of the 5'-triphosphate is correlated with efficient 3'-5' reverse polymerization. The intrinsic rate of removal of diphosphate from the G-1 residue of base-paired tRNAHis substrates is slow. Rates of diphosphate removal depend on the identity of the NTP included in reactions. The GTP 3'-OH is required to stimulate diphosphate removal
-
-
?
additional information
?
-
-
retention of the 5'-triphosphate is correlated with efficient 3'-5' reverse polymerization. The intrinsic rate of removal of diphosphate from the G-1 residue of base-paired tRNAHis substrates is slow. Rates of diphosphate removal depend on the identity of the NTP included in reactions. The GTP 3'-OH is required to stimulate diphosphate removal
-
-
?
additional information
?
-
the enzyme performs 3' to 5' or reverse polymerization
-
-
-
additional information
?
-
the GTP molecule is clearly bound at the active site, coordinating with two magnesium ions
-
-
-
additional information
?
-
-
the GTP molecule is clearly bound at the active site, coordinating with two magnesium ions
-
-
-
additional information
?
-
tRNAHis guanylyltransferase clearly prefers a substrate carrying a CCA terminus. The 3'-CCA triplet and a discriminator position A73 act as positive elements for G-1 incorporation, ensuring the fidelity of G-1 addition. On the substrate lacking the CCA-end (tRNAHisDELTACCA), the enzyme is less efficient
-
-
-
additional information
?
-
-
tRNAHis guanylyltransferase clearly prefers a substrate carrying a CCA terminus. The 3'-CCA triplet and a discriminator position A73 act as positive elements for G-1 incorporation, ensuring the fidelity of G-1 addition. On the substrate lacking the CCA-end (tRNAHisDELTACCA), the enzyme is less efficient
-
-
-
additional information
?
-
-
the enzyme also catalyzes 3'-5' extension of a polynucleotude chain by multiple nucleotides. Unlike the addition of G(-1), the reverse polymerisation activity is template-dependent, recognizing G*C WatsonCrick base pairs. Moreover, reverse polymerization is not specific for tRNAHis or for starting at the (-1) position of tRNA, provided that the activated (triphosphorylated) form of the tRNA substrate is used in the assays
-
-
?
NATURAL SUBSTRATE
NATURAL 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
p-tRNAHis + ATP + GTP
pppG-p-tRNAHis + AMP + diphosphate
the extra nucleotide in position -1 of mitochondrial and eukaryotic cytoplasmic tRNAHis molecules is added posttranscriptionally to the 5' end of the tRNA by a histidine-tRNA specific guanylyltransferase
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
-
-
-
?
p-tRNAHis + ATP + GTP
pppGp-tRNAHis + AMP + diphosphate
the G(-1) residue is both necessary and sufficient for aminoacylation of tRNA by histidyl-tRNA synthetase in vitro and is required for aminoacylation in vivo
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
-
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
overall reaction
-
-
?
p-tRNAHis + ATP + GTP + H2O
pGp-tRNAHis + AMP + 2 diphosphate
-
overall reaction
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
ATP
eukaryotic Thg1 enzymes require ATP for substrate activation
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Co2+
the requirement for a divalent metal ion can be satisfied by Mg2+, Mn2+, and to a lesser extent by Co2+ and Zn2+. Activity in presence of 10 mM Co2+ is 13% compared to the activity in presence of 10 mM Mg2+
Mg2+
Mn2+
the requirement for a divalent metal ion can be satisfied by Mg2+, Mn2+, and to a lesser extent by Co2+ and Zn2+. Activity in presence of 10 mM Mn2+ is 70% compared to the activity in presence of 10 mM Mg2+
Zn2+
the requirement for a divalent metal ion can be satisfied by Mg2+, Mn2+, and to a lesser extent by Co2+ and Zn2+. Activity in presence of 10 mM Zn2+ is 15% compared to the activity in presence of 10 mM Mg2+
Mg2+
the requirement for a divalent metal ion can be satisfied by Mg2+, Mn2+, and to a lesser extent by Co2+ and Zn2+. Maximal activity by 10 mM Mg2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
5,5'-dithiobis(2-nitrobenzoic)acid
progressive loss of activity with increasing amounts of DTNB. Thus, sulfhydryl groups are involved in maintaining the active state of the enzyme or are involved in the mechanism
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
additional information
analysis of G-1 incorporation on tRNAHis A73C variants with and without 3'-CCA-end shows significant preference of Thg1 for tRNAHis containing the 3'-CCA, both in terms of rate and maximal amount of product formed in the reactions, kinetic analysis
-
additional information
additional information
-
analysis of G-1 incorporation on tRNAHis A73C variants with and without 3'-CCA-end shows significant preference of Thg1 for tRNAHis containing the 3'-CCA, both in terms of rate and maximal amount of product formed in the reactions, kinetic analysis
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
7.5
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
22
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
malfunction
metabolism
in eukaryotes including yeast, both 3'-CCA and 5'-G-1 are added posttranscriptionally by tRNA nucleotidyltransferase and tRNAHis guanylyltransferase, respectively. These two cytosolic enzymes might compete for the same tRNA, but tRNAHis guanylyltransferase clearly prefers a substrate carrying a CCA terminus. Thus, although many tRNA maturation steps can occur in a rather random order, pathway where CCA-addition precedes G-1 incorporation is likely in Saccharomyces cerevisiae. The 3'-CCA triplet and a discriminator position A73 act as positive elements for G-1 incorporation, ensuring the fidelity of G-1 addition. Sequential order of tRNAHis processing, overview. The enzymes do not compete for the substrate. Instead, the differing substrate preferences lead to a sequential order of nucleotide incorporation at 5'- and 3'-ends, resulting in a mature tRNAHis in the cytosol
physiological function
additional information
evolution
the active site moiety of ScThg1 is highly conserved among members of the Thg1 family, comparisons of structure ana binding modes, overview
evolution
the archaeal type Thg1, which includes most archaeal and bacterial Thg1 enzymes, is phylogenetically distant from eukaryotic Thg1
evolution
the tRNAHis guanylyltransferase (Thg1) superfamily includes enzymes that are found in all three domains of life that all share the common ability to catalyze the 3' to 5' synthesis of nucleic acids. This catalytic activity is the reverse of all other known DNA and RNA polymerases. Forward 5' to 3' and reverse 3' to 5' polymerization are mechanistically similar. In bacteria and many archaea, the G-1 residue is genomically encoded and transcribed in the precursor tRNA transcript, and subsequent cleavage of the 5' leader sequence by ribonuclease P (RNase P) yields a mature tRNAHis with its identity-establishing G-1 element (tRNAHisG-1). Different pathways to establish tRNAHis identity, overview. Thg1-like proteins function in tRNA repair, TLPs in bacteria and archaea, structural comparison of Thg1 and TLPs. TLPs exhibit broader RNA recognition properties than Thg1 homologues
evolution
tRNAHis guanylyltransferase (Thg1) specifies eukaryotic tRNAHis identity by catalyzing a 3' to 5' non-Watson Crick (WC) addition of guanosine to the 5'-end of tRNAHis. Thg1 family enzymes in Archaea and Bacteria, called Thg1-like-proteins (TLPs), catalyze a similar but distinct 3' to 5' addition in an exclusively WC-dependent manner
malfunction
a thg1DELTA strain is only viable in the presence of a functional version of the G-1 addition enzyme supplied on a covering plasmid. Using this system, different archaeal TLPs are not able to support growth of the thg1DELTA strain, and only strains that also simultaneously express a mutant form of tRNAHis (C73-tRNAHis), enabling WC-dependent addition of G-1, are viable. BtTLP is able to carry out the essential activity ScThg1 in vivo in Saccharomyces cerevisiae, but this apparent function contradicts the strong preference for WC base-paired nucleotide addition exhibited by BtTLP in vitro. BtTLP catalyzes its biochemically preferred WC-dependent reaction, possibly adding U-1 to this A73-containing tRNA, mechanism, overview
malfunction
identification of residues in yeast Thg1 that play a part in preventing extended polymerization. Mutation of these residues with alanine results in extended reverse polymerization
physiological function
the diphosphate removal activity under all conditions preferentially acts upon tRNAs containing a U-1:A73 terminating base pair. Although Thg1 can add UTP to create a U:A base pair, it efficiently removes the activated 5'-end from this added nucleotide and effectively terminates subsequent addition reactions. Thg1 exhibits 3'-5' polymerization with some tRNA substrates, but addition of a nucleotide at the -1 position is the preferred reaction
physiological function
the tRNAHis guanylyltransferase posttranscriptionally adds a 5'-G-1 from GTP to yeast tRNAHis. The presence of this single G at the -1 position is critical for recognition of tRNAHis by the histidyl-tRNA synthetase (HisRS). Thg1 adds a single G-1 residue across from an A73 discriminator nucleotide, forming an unconventional non-Watson Crick base pair. This unusual 3'-5' nucleotidyltransferase protein is a member of a larger family of Watson Crick-dependent 3'-5' polymerase enzymes that share structural similarity with canonical 5'-3' DNA and RNA polymerases
physiological function
tRNAHis guanylyltransferase (Thg1) has unique reverse (3'-5') polymerase activity occurring in all three domains of life. Most eukaryotic Thg1 homologs are essential genes involved in tRNAHis maturation. These enzymes normally catalyze a single 5' guanylation of tRNAHis lacking the essential G-1 identity element required for aminoacylation. G-1 is the critical identity element that histidyl-tRNA synthetase (HisRS) uses to recognize its cognate tRNA, enabling HisRS to differentiate tRNAHis from the pool of other cellular tRNAs
physiological function
tRNAHis guanylyltransferase (Thg1) post-transcriptionally adds G-1. Histidyl-tRNA synthetase (HisRS) recognizes the Thg1-incorporated G-1 for the accurate histidylation of tRNAHis in eukaryotes. Role of 3' to 5' reverse RNA polymerization in tRNA fidelity and repair. G-1 is a tRNAHis identity element
physiological function
tRNAHis guanylyltransferase (Thg1) specifies eukaryotic tRNAHis identity by catalyzing a 3' to 5' non-Watson Crick (WC) addition of guanosine to the 5'-end of tRNAHis. Thg1 family enzymes in Archaea and Bacteria, called Thg1-like-proteins (TLPs), catalyze a similar but distinct 3' to 5' addition in an exclusively WC-dependent manner. Essential nature of ScThg1
additional information
identification of residues in yeast Thg1 that play a part in preventing extended polymerization. Mutation of these residues with alanine results in extended reverse polymerization
additional information
-
identification of residues in yeast Thg1 that play a part in preventing extended polymerization. Mutation of these residues with alanine results in extended reverse polymerization
additional information
molecular basis for tRNA recognition, and catalytic mechanism, overview. Molecular basis for non-Watson-Crick G-1 addition: tRNA activation, nucleotidyl transfer, diphosphate removal, and maintenance of tRNAHis fidelity
additional information
two flexible protomers at the potential binding site (PBS) for tRNAHis are observed, the PBS of the tetramer could also be one of the sites for interaction with partner proteins
additional information
-
two flexible protomers at the potential binding site (PBS) for tRNAHis are observed, the PBS of the tetramer could also be one of the sites for interaction with partner proteins
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
tetramer
additional information
eukaryotic Thg1 enzymes show monomeric Thg1 that is composed of a six-strand antiparallel beta-sheet flanked by three or four alpha-helices on each side, along with a protruding long arm composed of two antiparallel beta-strands. Structure-function analyses of Thg1 and TLP enzymes, overview
tetramer
4 * 27800, about, sequence calculation and crystal structure
tetramer
crystal structures of eukaryotic Thg1 enzymes showed that monomeric Thg1 is composed of a six-strand antiparallel beta-sheet flanked by three or four alpha-helices on each side, along with a protruding long arm composed of two antiparallel beta-strands. Each monomer forms a dimer, mediated by salt bridge formation and hydrogen bonding between the beta-sheet and an alpha-helix. Alanine scanning mutagenesis in Saccharomyces cerevisiae Thg1 (ScThg1) of conserved residues responsible for mediating dimer formation has shown strongly diminished G-1 addition activity. Dimer-of-dimer formation stems from initial dimer formation., the alpha-helix of the N-terminus in monomer A interacts with the nucleotide binding site of monomer B and ultimately forms an intertwined N-terminus segment, which again interacts with a symmetrical dimer to form its active tetrameric form
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified GTP-bound Thg1, hanging drop vapor diffusion method, mixing of 0.001 ml of 7 mg/ml protein in 20 mM Tris, pH 8.0, 500 mM NaCl, and 5 mM DTT, with 0.001 ml of reservoir solution consisting of 0.1 M 2-(N-morpholino)ethanesulfonic acid, pH 6.0, and 0.9 M ammonium sulfate, equilibration against 0.5 ml of reservoir solution, X-ray diffraction structure determination and analysis at 3.0 A resolution, molecular replacement method, modelling
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D153A
poorly expressed. D153A is stable, but its expression is somehow toxic to Escherichia coli. 312% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
D47A
54.7% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
D68A
59% of wild-type G(-1) addition activity. The mutation causes a dramatic decrease in the rigorous specificity of Thg1 for tRNAHis. This single alteration enhances the kcat/KM for ppp-tRNAPhe by nearly 100-fold relative to that of wild-type Thg1. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
D77A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
E13A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
E75A
105% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
E78A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
G74A
44.4% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
H155A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
K190A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
K211A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
K44A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
K96A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
N161A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
N201A
less than 0.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
N46A
22.6% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
P45A
62.3% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
R133A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
R150A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
R27A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
R93A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
W113A/I156V
40.1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
Y160A
0.1%-1% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
Y202A
poorly expressed, The Y202A variant is present at very low levels in soluble crude extracts and yields nearly undetectable levels of protein after purification, suggesting that this alteration leads to lack of overall stability
Y40A
42.9% of wild-type G(-1) addition activity. G(-) i.e. the extra nucleotide posttranscriptionally added in position (-1) of tRNA
additional information
additional information
Saccharomyces cerevisiae tRNAHis transcripts with and without G-1 are generated, carrying a C73 discriminator instead of the wild-type A73 position (tRNAHisDELTAG-1 A73C and tRNAHis+G-1 A73C). Due to the additional base pair G-1/C73, tRNAHis+G-1 A73C carries an extended acceptor stem with a base-paired discriminator position. This situation does not affect CCA-addition catalyzed by the CCA-adding enzyme and results in a similarly efficient CCA incorporation to tRNAHis+G-1 A73C compared to tRNAHisDELTAG-1 A73C. Both tRNAHis substrates with a cytosine at the discriminator position are readily accepted as substrates for CCA-addition, showing comparable band patterns like the wild-type tRNAHis containing an A73. Analysis of G-1 incorporation on tRNAHis A73C variants with and without 3'-CCA-end shows significant preference of Thg1 for tRNAHis containing the 3'-CCA, both in terms of rate and maximal amount of product formed in the reactions, kinetic analysis
additional information
-
Saccharomyces cerevisiae tRNAHis transcripts with and without G-1 are generated, carrying a C73 discriminator instead of the wild-type A73 position (tRNAHisDELTAG-1 A73C and tRNAHis+G-1 A73C). Due to the additional base pair G-1/C73, tRNAHis+G-1 A73C carries an extended acceptor stem with a base-paired discriminator position. This situation does not affect CCA-addition catalyzed by the CCA-adding enzyme and results in a similarly efficient CCA incorporation to tRNAHis+G-1 A73C compared to tRNAHisDELTAG-1 A73C. Both tRNAHis substrates with a cytosine at the discriminator position are readily accepted as substrates for CCA-addition, showing comparable band patterns like the wild-type tRNAHis containing an A73. Analysis of G-1 incorporation on tRNAHis A73C variants with and without 3'-CCA-end shows significant preference of Thg1 for tRNAHis containing the 3'-CCA, both in terms of rate and maximal amount of product formed in the reactions, kinetic analysis
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
the enzyme shows a drastically shortened half-life in the absence of divalent metal ions
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant C-terminally His6-tagged enzyme from Escherichia coli strain Rosetta(DE3) by nickel affinity chromatography, ultrafiltration, and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene thg1, recombinant expression of C-terminally His6-tagged enzyme in Escherichia coli strain Rosetta(DE3)
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Jackman, J.E.; Phizicky, E.M.
Identification of critical residues for G-1 addition and substrate recognition by tRNA(His) guanylyltransferase
Biochemistry
47
4817-4825
2008
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Rice, T.S.; Ding, M.; Pederson, D.S.; Heintz, N.H.
The highly conserved tRNAHis guanylyltransferase Thg1p interacts with the origin recognition complex and is required for the G2/M phase transition in the yeast Saccharomyces cerevisiae
Eukaryot. Cell
4
832-835
2005
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Gu, W.; Jackman, J.E.; Lohan, A.J.; Gray, M.W.; Phizicky, E.M.
tRNAHis maturation: an essential yeast protein catalyzes addition of a guanine nucleotide to the 5' end of tRNAHis
Genes Dev.
17
2889-2901
2003
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Pande, S.; Jahn, D.; Sll, D.
Histidine tRNA guanylyltransferase from Saccharomyces cerevisiae. I. Purification and physical properties
J. Biol. Chem.
266
22826-22831
1991
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Jahn, D.; Pande, S.
Histidine tRNA guanylyltransferase from Saccharomyces cerevisiae. II. Catalytic mechanism
J. Biol. Chem.
266
22832-22836
1991
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Gu, W.; Hurto, R.L.; Hopper, A.K.; Grayhack, E.J.; Phizicky, E.M.
Depletion of Saccharomyces cerevisiae tRNA(His) guanylyltransferase Thg1p leads to uncharged tRNAHis with additional m(5)C
Mol. Cell. Biol.
25
8191-8201
2005
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Jackman, J.E.; Phizicky, E.M.
tRNAHis guanylyltransferase catalyzes a 3'-5' polymerization reaction that is distinct from G-1 addition
Proc. Natl. Acad. Sci. USA
103
8640-8645
2006
Saccharomyces cerevisiae
Jackman, J.E.; Phizicky, E.M.
tRNAHis guanylyltransferase adds G-1 to the 5' end of tRNAHis by recognition of the anticodon, one of several features unexpectedly shared with tRNA synthetases
RNA
12
1007-1014
2006
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Preston, M.A.; Phizicky, E.M.
The requirement for the highly conserved G-1 residue of Saccharomyces cerevisiae tRNAHis can be circumvented by overexpression of tRNAHis and its synthetase
RNA
16
1068-1077
2010
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Smith, B.A.; Jackman, J.E.
Saccharomyces cerevisiae Thg1 uses 5-pyrophosphate removal to control addition of nucleotides to tRNA(His.)
Biochemistry
53
1380-1391
2014
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae
Lee, K.; Lee, E.H.; Son, J.; Hwang, K.Y.
Crystal structure of tRNAHis guanylyltransferase from Saccharomyces cerevisiae
Biochem. Biophys. Res. Commun.
490
400-405
2017
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 2045083 (P53215)
Dodbele, S.; Moreland, B.; Gardner, S.; Bundschuh, R.; Jackman, J.
5'-End sequencing in Saccharomycesxa0cerevisiae offers new insights into 5'-ends of tRNAHis and snoRNAs
FEBS Lett.
593
971-981
2019
Bacillus thuringiensis serovar israelensis (Q3F0V8), Bacillus thuringiensis serovar israelensis ATCC 35646 (Q3F0V8), Dictyostelium discoideum (Q54WD4), Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae ATCC 204508 (P53215)
Chen, A.; Jayasinghe, M.; Chung, C.; Rao, B.; Kenana, R.; Heinemann, I.; Jackman, J.
The role of 3' to 5' reverse RNA polymerization in tRNA fidelity and repair
Genes (Basel)
10
250
2019
Bacillus thuringiensis (A0A2A8WD07), Bacillus thuringiensis T01-328 (A0A2A8WD07), Candida albicans (A0A1D8PQL3), Candida albicans ATCC MYA-2876 (A0A1D8PQL3), Dictyostelium discoideum (Q54E29), Dictyostelium discoideum (Q54HW0), Dictyostelium discoideum (Q54WD4), Dictyostelium discoideum (Q86IE7), Homo sapiens (Q9NWX6), Ignicoccus hospitalis (A8A9R6), Ignicoccus hospitalis DSM 18386 (A8A9R6), Ignicoccus hospitalis JCM 14125 (A8A9R6), Ignicoccus hospitalis KIN4/I (A8A9R6), Methanopyrus kandleri (Q8TZ46), Methanopyrus kandleri DSM 6324 (Q8TZ46), Methanopyrus kandleri JCM 9639 (Q8TZ46), Methanopyrus kandleri NBRC 100938 (Q8TZ46), Methanosarcina acetivorans (A0A1C7D1G9), Methanosarcina barkeri (Q46BQ4), Methanosarcina barkeri DSM 804 (Q46BQ4), Methanosarcina barkeri Fusaro (Q46BQ4), Methanothermobacter thermautotrophicus, Myxococcus xanthus (Q1CZS0), Myxococcus xanthus DK 1622 (Q1CZS0), no activity in Acanthamoeba castellanii, no activity in Trypanosoma brucei, Pyrobaculum aerophilum (Q8ZY97), Pyrobaculum aerophilum ATCC 51768 (Q8ZY97), Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae ATCC 204508 (P53215)
Poehler, M.T.; Roach, T.M.; Betat, H.; Jackman, J.E.; Moerl, M.
A temporal order in 5'-and 3'-processing of eukaryotic tRNAHis
Int. J. Mol. Sci.
20
1384
2019
Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (P53215)
Desai, R.; Kim, K.; Buechsenschuetz, H.C.; Chen, A.W.; Bi, Y.; Mann, M.R.; Turk, M.A.; Chung, C.Z.; Heinemann, I.U.
Minimal requirements for reverse polymerization and tRNA repair by tRNAHis guanylyltransferase
RNA Biol.
15
614-622
2018
Ignicoccus hospitalis (A8A9R6), Ignicoccus hospitalis, Methanosarcina acetivorans (A0A1C7D1G9), Methanosarcina acetivorans, Pyrobaculum aerophilum (Q8ZY97), Pyrobaculum aerophilum, Pyrobaculum aerophilum ATCC 51768 (Q8ZY97), Pyrobaculum aerophilum DSM 7523 (Q8ZY97), Pyrobaculum aerophilum IM2 (Q8ZY97), Pyrobaculum aerophilum JCM 9630 (Q8ZY97), Pyrobaculum aerophilum NBRC 100827 (Q8ZY97), Saccharomyces cerevisiae (P53215), Saccharomyces cerevisiae, Saccharomyces cerevisiae ATCC 204508 (P53215)
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