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2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP-ribose 1'',2''-phosphate + nicotinamide
-
-
-
?
18-mer 2' phosphorylated RNA
?
-
pACAAGACUCUAA-(2-phosphate)AUCUUG
-
-
?
18-mer 2' phosphorylated RNA + NAD+
?
-
ACAAGACUGUAA-(2-phosphate)AUCUUG
-
-
?
2'-phospho-[ligated tRNA]
?
-
-
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP-ribose 1'',2''-phosphate + nicotinamide
-
-
-
-
?
3-mer 2' phosphorylated RNA
?
-
pApA-(2-phosphate)pA, pApU-(2-phosphate)pA, pUpU-(2-phosphate)pU, pApApA-(2-phosphate),pApApA-(2-phosphate)p, pApApA-(2-phosphate)1OCH3
-
-
?
3-mer 2' phosphorylated RNA + NAD+
?
-
AU-(2-phosphate)U
-
-
?
8-mer 2' phosphorylated RNA
?
-
pGUAA-(2-phosphate)AUCU
-
-
?
8-mer 2' phosphorylated RNA + NAD+
?
-
GUAA-(2-phosphate)AUCU
-
-
?
NAD+ + trinucleotide substrate
ADP-ribosylated RNA
-
requires ca. 0.0004 mM Tpt1 K69A/R71S, only 0.0000003 mM Tpt1p is required for similar or greater product formation
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
-
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
minimal substrate is a dinucleotide containing an internal 2'-phosphate
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
specificity for substrates bearing an internal 2'-phosphate
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
Tpt1 can catalyze the conversion of the KptA-generated intermediate to both product and the original substrate
-
-
?
2'-phospho-[ligated tRNA] + NAD+
mature tRNA + ADP ribose-1'',2''-phosphate + nicotinamide + H2O
-
tRNA_Leu_UAG, tRNA_SER, tRNA_LEU_CAA, tRNA_TYR, tRNA_LYS, tRNA_PRO1, tRNA_PRO2, tRNA_PHE, unidentified tRNA
-
-
?
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C85R
-
no accumulation of the reaction intermediate
D34A
-
no accumulation of the reaction intermediate
F72A
-
temperature sensitive
H117A-T119A
-
lethal in vivo
H140A
-
no accumulation of the reaction intermediate
H140A/H142A
-
no accumulation of the reaction intermediate
H140A/V141A/H142A
-
no accumulation of the reaction intermediate
I167L/D168K
-
no accumulation of the reaction intermediate
K16A
-
colony size indistinguishable from that of wild-type TPT1 strain
K69A
-
colony size indistinguishable from that of wild-type TPT1 strain
K69A/R71S
-
this mutant produces a substantial amount of the reaction intermediate
M136G
-
no accumulation of the reaction intermediate
N185A
-
no accumulation of the reaction intermediate
Q88A/H90A/S91A
-
no accumulation of the reaction intermediate
Q88Y
-
no accumulation of the reaction intermediate
R138K
-
colony size indistinguishable from that of wild-type TPT1 strain
R158A
-
colony size indistinguishable from that of wild-type TPT1 strain
R158H
-
no accumulation of the reaction intermediate
R23A/H24A
-
no accumulation of the reaction intermediate
R71Q
-
grows slower than wild-type, temperature sensitive
S155A
-
no accumulation of the reaction intermediate
S15A/K16A
-
no accumulation of the reaction intermediate
S183A
-
no accumulation of the reaction intermediate
T119A
-
colony size indistinguishable from that of wild-type TPT1 strain
Y38A
-
colony size indistinguishable from that of wild-type TPT1 strain
additional information
generation and analysis of diverse mutant strains derived from Saccharomyces cerevisiae strain W303. The essential functions of TRL1 and TPT1 in budding yeast are bypassed by expressing prespliced, intronless versions of the 10 normally intron-containing tRNAs, indicating this repair pathway does not have additional essential functions. Expression of intronless tRNAs fails to rescue the growth of cells with deletions in components of the SEN complex, implying an additional essential role for the splicing endonuclease. The trl1DELTA and tpt1DELTA mutants accumulate tRNA and HAC1 splicing intermediates indicative of RNA repair defects and are hypersensitive to drugs that inhibit translation. tpt1DELTA mutants grow in the presence of tunicamycin despite reduced accumulation of spliced HAC1 mRNA, while failure to induce the unfolded protein response occurs in trl1DELTA cells grown with tunicamycin is lethal owing to their inability to ligate HAC1 after its cleavage by Ire1. Tpt1DELTA phenotype, overview
H117A
-
colony size indistinguishable from that of wild-type TPT1 strain
H117A
-
no accumulation of the reaction intermediate
H142A
-
colony size indistinguishable from that of wild-type TPT1 strain
H142A
-
no accumulation of the reaction intermediate
H90A
-
colony size indistinguishable from that of wild-type TPT1 strain
H90A
-
no accumulation of the reaction intermediate
R138A
-
lethal in vivo
R138A
-
no accumulation of the reaction intermediate
S91A
-
colony size indistinguishable from that of wild-type TPT1 strain
S91A
-
no accumulation of the reaction intermediate
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Culver, G.M.; McCraith, S.M.; Consaul, S.A.; Stanford, D.R.; Phizicky, E.M.
A 2'-phosphotransferase implicated in tRNA splicing is essential in Saccharomyces cerevisiae
J. Biol. Chem.
272
13203-13210
1997
Escherichia coli, Homo sapiens, Mus musculus, Saccharomyces cerevisiae, Saccharomyces cerevisiae (Q12272), Schizosaccharomyces pombe
brenda
Steiger, M.A.; Kierzek, R.; Turner, D.H.; Phizicky, E.M.
Substrate recognition by a yeast 2'-phosphotransferase involved in tRNA splicing and by its Escherichia coli homolog
Biochemistry
40
14098-14105
2001
Escherichia coli, Saccharomyces cerevisiae
brenda
Hu, Q.D.; Lu, H.; Huo, K.; Ying, K.; Li, J.; Xie, Y.; Mao, Y.; Li, Y.Y.
A human homolog of the yeast gene encoding tRNA 2'-phosphotransferase: cloning, characterization and complementation analysis
Cell. Mol. Life Sci.
60
1725-1732
2003
Arabidopsis thaliana, Archaeoglobus fulgidus, Clostridium perfringens, Escherichia coli, Fusobacterium nucleatum, Homo sapiens, Homo sapiens (Q86TN4), Methanopyrus kandleri, Mus musculus, Rattus norvegicus, Saccharomyces cerevisiae, Schizosaccharomyces pombe, Streptomyces coelicolor
brenda
Mc Craith, S.M.; Phizicky, E.M.
An enzyme from Saccharomyces cerevisiae uses NAD+ to transfer the splice junction 2'-phosphate from ligated tRNA to an acceptor molecule
J. Biol. Chem.
266
11986-11992
1991
Saccharomyces cerevisiae
brenda
Zillman, M.; Gorovsky, M.A.; Phizicky, E.M.
HeLa cells contain a 2'-phosphate-specific phosphotransferase similar to a yeast enzyme implicated in tRNA splicing
J. Biol. Chem.
267
10289-10294
1992
Homo sapiens, Saccharomyces cerevisiae
brenda
Spinelli, S.L.; Kierzek, R.; Turner, D.H.; Phizicky, E.M.
Transient ADP-ribosylation of a 2'-phosphate implicated in its removal from ligated tRNA during splicing in yeast
J. Biol. Chem.
274
2637-2644
1999
Saccharomyces cerevisiae, Escherichia coli, Mus musculus
brenda
Kato-Murayama, M.; Bessho, Y.; Shirouzu, M.; Yokoyama, S.
Crystal structure of the RNA 2'-phosphotransferase from Aeropyrum pernix K1
J. Mol. Biol.
348
295-305
2005
Aeropyrum pernix, Arabidopsis thaliana, Archaeoglobus fulgidus, Escherichia coli, Homo sapiens, Methanosarcina mazei, Pseudomonas aeruginosa, Pyrococcus horikoshii, Saccharomyces cerevisiae
brenda
Kierzek, R.; Steiger, M.A.; Spinelli, S.L.; Turner, D.H.; Phizicky, E.M.
The chemical synthesis of oligoribonucleotides with selectively placed 2'-O-phosphates
Nucleosides Nucleotides Nucleic Acids
19
917-933
2000
Saccharomyces cerevisiae
brenda
Spinelli, S.L.; Malik, H.S.; Consaul, S.A.; Phizicky, E.M.
A functional homolog of a yeast tRNA splicing enzyme is conserved in higher eukaryotes and in Escherichia coli
Proc. Natl. Acad. Sci. USA
95
14136-14141
1998
Arabidopsis thaliana, Archaeoglobus fulgidus, Candida albicans, Escherichia coli, Mus musculus, no activity in Bacillus subtilis, no activity in Haemophilus influenzae, no activity in Helicobacter pylori, no activity in Mycoplasma genitalium, Pseudomonas aeruginosa, Pyrococcus horikoshii, Saccharomyces cerevisiae, Schizosaccharomyces pombe
brenda
Sawaya, R.; Schwer, B.; Shuman, S.
Structure-function analysis of the yeast NAD+-dependent tRNA 2'-phosphotransferase Tpt1
RNA
11
107-113
2005
Aeropyrum pernix, Archaeoglobus fulgidus, Clostridium perfringens, Drosophila melanogaster, Escherichia coli, Homo sapiens, Leishmania major, Nostoc punctiforme, Pyrococcus horikoshii, Saccharomyces cerevisiae, Schizosaccharomyces pombe, Trypanosoma cruzi
brenda
Steiger, M.A.; Jackman, J.E.; Phizicky, E.M.
Analysis of 2'-phosphotransferase (Tpt1p) from Saccharomyces cerevisiae: evidence for a conserved two-step reaction mechanism
RNA
11
99-106
2005
Saccharomyces cerevisiae
brenda
Spinelli, S.L.; Consaul, S.A.; Phizicky, E.M.
A conditional lethal yeast phosphotransferase (tpt1) mutant accumulates tRNAs with a 2'-phosphate and an undermodified base at the splice junction
RNA
3
1388-1400
1997
Escherichia coli, Saccharomyces cerevisiae
brenda
Sorci, L.; Ruggieri, S.; Raffaelli, N.
NAD homeostasis in the bacterial response to DNA/RNA damage
DNA Repair
23
17-26
2014
Agrobacterium tumefaciens (Q8U9Z2), Agrobacterium tumefaciens C58 / ATCC 33970 (Q8U9Z2), Brachyspira hyodysenteriae (A0A3B6VB17), Brachyspira hyodysenteriae ATCC 49526 (A0A3B6VB17), Catenulispora acidiphila (C7QAQ2), Deinococcus deserti (C1CZ01), Deinococcus deserti DSM 17065 (C1CZ01), Deinococcus gobiensis (H8GTD5), Deinococcus gobiensis DSM 21396 (H8GTD5), Deinococcus radiodurans (Q9RRR1), Deinococcus radiodurans ATCC 13939 (Q9RRR1), Delftia acidovorans (A9C0S5), Delftia acidovorans DSM 14801 (A9C0S5), Flavobacterium johnsoniae (A5FLZ4), Fusobacterium nucleatum subsp. nucleatum (Q8R5N7), Fusobacterium nucleatum subsp. nucleatum ATCC 25586 / JCM14847 (Q8R5N7), Herpetosiphon aurantiacus (A9B356), Herpetosiphon aurantiacus DSM 785 (A9B356), Saccharomyces cerevisiae, Sebaldella termitidis (D1APE3), Sebaldella termitidis ATCC 33386 (D1APE3)
brenda
Munir, A.; Banerjee, A.; Shuman, S.
NAD+-dependent synthesis of a 5-phospho-ADP-ribosylated RNA/DNA cap by RNA 2-phosphotransferase Tpt1
Nucleic Acids Res.
46
9617-9624
2018
Acetivibrio thermocellus (A3DJX6), Acetivibrio thermocellus ATCC 27405 (A3DJX6), Acetivibrio thermocellus DSM 1237 (A3DJX6), Acetivibrio thermocellus JCM 9322 (A3DJX6), Acetivibrio thermocellus NBRC 103400 (A3DJX6), Acetivibrio thermocellus NCIMB 10682 (A3DJX6), Acetivibrio thermocellus NRRL B-4536 (A3DJX6), Acetivibrio thermocellus VPI 7372 (A3DJX6), Aeropyrum pernix (Q9YFP5), Aeropyrum pernix, Aeropyrum pernix ATCC 700893 (Q9YFP5), Aeropyrum pernix DSM 11879 (Q9YFP5), Aeropyrum pernix JCM 9820 (Q9YFP5), Aeropyrum pernix NBRC 100138 (Q9YFP5), Archaeoglobus fulgidus (O29841), Archaeoglobus fulgidus ATCC 49558 (O29841), Archaeoglobus fulgidus JCM 9628 (O29841), Archaeoglobus fulgidus NBRC 100126 (O29841), Archaeoglobus fulgidus VC-16 (O29841), Homo sapiens (Q86TN4), Pyrococcus horikoshii (O57899), Pyrococcus horikoshii ATCC 700860 (O57899), Pyrococcus horikoshii DSM 12428 (O57899), Pyrococcus horikoshii JCM 9974 (O57899), Pyrococcus horikoshii NBRC 100139 (O57899), Pyrococcus horikoshii OT-3 (O57899), Runella slithyformis, Saccharomyces cerevisiae (Q12272), Thermochaetoides thermophila (G0S5Z5), Thermochaetoides thermophila CBS 144.50 (G0S5Z5), Thermochaetoides thermophila DSM 1495 (G0S5Z5), Thermochaetoides thermophila IMI 039719 (G0S5Z5)
brenda
Cherry, P.D.; White, L.K.; York, K.; Hesselberth, J.R.
Genetic bypass of essential RNA repair enzymes in budding yeast
RNA
24
313-323
2018
Saccharomyces cerevisiae (Q12272)
brenda