The Experts below are selected from a list of 705 Experts worldwide ranked by ideXlab platform
James A. Mccloskey - One of the best experts on this subject based on the ideXlab platform.
-
POSTTRANSCRIPTIONAL MODIFICATION OF TRNA IN PSYCHROPHILIC BACTERIA
Journal of bacteriology, 1997Co-Authors: Joseph J. Dalluge, Richard Y. Morita, Tetsuo Hamamoto, Koki Horikoshi, Karl O. Stetter, James A. MccloskeyAbstract:Posttranscriptional modification in tRNA is known to play a multiplicity of functional roles, including maintenance of tertiary structure and cellular adaptation to environmental factors such as temperature. Nucleoside modification has been studied in unfractionated tRNA from three psychrophilic bacteria (ANT-300 and Vibrio sp. strains 5710 and 29-6) and one psychrotrophic bacterium (Lactobacillus bavaricus). Based on analysis of total enzymatic hydrolysates by liquid chromatography-mass spectrometry, unprecedented low amounts of modification were found in the psychrophiles, particularly from the standpoint of structural diversity of modifications observed. Thirteen to 15 different forms of posttranscriptional modification were found in the psychrophiles, and 10 were found in L. bavaricus, compared with approximately 29 known to occur in bacterial mesophiles and 24 to 31 known to occur in the archaeal hyperthermophiles. The four most abundant modified nucleosides in tRNA from each organism were Dihydrouridine, pseudouridine, 7-methylguanosine, and 5-methyluridine. The molar abundances of the latter three nucleosides were comparable to those found in tRNA from Escherichia coli. By contrast, the high levels of Dihydrouridine observed in all three psychrophiles are unprecedented for any organism in any of the three phylogenetic domains. tRNA from these organisms contains 40 to 70% more Dihydrouridine, on average, than that of the mesophile E. coli or the psychrotroph L. bavaricus. This finding supports the concept that a functional role for Dihydrouridine is in maintenance of conformational flexibility of RNA, especially important to organisms growing under conditions where the dynamics of thermal motion are severely compromised. This is in contrast to the role of modifications contained in RNA from thermophiles, which is to reduce regional RNA flexibility and provide structural stability to RNA for adaptation to high temperature.
-
Quantitative measurement of Dihydrouridine in RNA using isotope dilution liquid chromatography–mass spectrometry (LC/MS)
1996Co-Authors: Joseph J. Dalluge, Takeshi Hashizume, James A. MccloskeyAbstract:A method has been developed for the microscale determination of 5,6-Dihydrouridine, the most common post-transcriptional modification in bacterial and euka-ryotic tRNA. The method is based on stable isotope dilution liquid chromatography–mass spectrometry (LC/ MS) using [1,3-15N2]Dihydrouridine and [1,3-15N2]uridine as internal standards. RNA samples were enzymatically digested to nucleosides before addition of the internal standards and subsequently analyzed by LC/MS with selected ion monitoring of protonated molecular ions of the labeled and unlabeled nucleosides. Sample quantities of ∼1 pmol tRNA and 5 pmol 23S rRNA were analyzed for mole % Dihydrouridine. Dihydrouridine content of Escherichia coli tRNASerVGA and tRNAThrGGU as controls were measured as 2.03 and 2.84 residues/ tRNA molecule, representing accuracies of 98 and 95%. Overall precision values for the analyses of E.coli tRNASerVGA and E.coli tRNAThrGGU, unfractionated tRNA from E.coli and 23S rRNA from E.coli were within the range 0.43–2.4%. The mole % Dihydrouridine in unfrac-tionated tRNA and 23S rRNA from E.coli were deter-mined as 1.79 and 0.0396%, corresponding to 1.4 and 1.1 residues/RNA molecule respectively
-
Conformational flexibility in RNA: the role of Dihydrouridine
1996Co-Authors: Joseph J. Dalluge, James A. Mccloskey, Takeshi Hashizume, Alan E. Sopchik, Darrell R. DavisAbstract:In order to further understand the structural role of the modified nucleoside Dihydrouridine in RNA the solution conformations of Dp and ApDpA were analyzed by one-and two-dimensional proton NMR spectroscopy and compared with those of the related uridine-containing compounds. The analyses indicate that Dihydrouridine significantly destabilizes the C3′-endo sugar con-formation associated with base stacked, ordered, A-type helical RNA. Equilibrium constants (Keq = [C2′-endo]/[C3′-endo]) for C2′-endo–C3′-endo interconversion at 25C for Dp, the 5′-terminal A of ApDpA and D in ApDpA are 2.08, 1.35 and 10.8 respectively. Stabilization of the C2′-endo form was shown to be enhanced at low temperature, indicating that C2′-endo is the thermo-dynamically favored conformation for Dihydrouridine. ∆H values show that for Dp the C2′-endo sugar conformation is stabilized by 1.5 kcal/mol compared with Up. This effect is amplified for D in the oligonucleotide ApDpA and propagated to the 5′-neighboring A, with stabilization of the C2′-endo form by 5.3 kcal/mol for D and 3.6 kcal/mol for the 5′-terminal A. Post-tran-scriptional formation of Dihydrouridine therefore represents a biological strategy opposite in effect to ribose methylation, 2-thiolation or pseudouridylation, all of which enhance regional stability through stabilization of the C3′-endo conformer. Dihydrouridine effectively promotes the C2′-endo sugar conformation, allowing for greater conformational flexibility and dynamic motion in regions of RNA where tertiary interactions and loop formation must be simultaneously accomodated
Joseph J. Dalluge - One of the best experts on this subject based on the ideXlab platform.
-
POSTTRANSCRIPTIONAL MODIFICATION OF TRNA IN PSYCHROPHILIC BACTERIA
Journal of bacteriology, 1997Co-Authors: Joseph J. Dalluge, Richard Y. Morita, Tetsuo Hamamoto, Koki Horikoshi, Karl O. Stetter, James A. MccloskeyAbstract:Posttranscriptional modification in tRNA is known to play a multiplicity of functional roles, including maintenance of tertiary structure and cellular adaptation to environmental factors such as temperature. Nucleoside modification has been studied in unfractionated tRNA from three psychrophilic bacteria (ANT-300 and Vibrio sp. strains 5710 and 29-6) and one psychrotrophic bacterium (Lactobacillus bavaricus). Based on analysis of total enzymatic hydrolysates by liquid chromatography-mass spectrometry, unprecedented low amounts of modification were found in the psychrophiles, particularly from the standpoint of structural diversity of modifications observed. Thirteen to 15 different forms of posttranscriptional modification were found in the psychrophiles, and 10 were found in L. bavaricus, compared with approximately 29 known to occur in bacterial mesophiles and 24 to 31 known to occur in the archaeal hyperthermophiles. The four most abundant modified nucleosides in tRNA from each organism were Dihydrouridine, pseudouridine, 7-methylguanosine, and 5-methyluridine. The molar abundances of the latter three nucleosides were comparable to those found in tRNA from Escherichia coli. By contrast, the high levels of Dihydrouridine observed in all three psychrophiles are unprecedented for any organism in any of the three phylogenetic domains. tRNA from these organisms contains 40 to 70% more Dihydrouridine, on average, than that of the mesophile E. coli or the psychrotroph L. bavaricus. This finding supports the concept that a functional role for Dihydrouridine is in maintenance of conformational flexibility of RNA, especially important to organisms growing under conditions where the dynamics of thermal motion are severely compromised. This is in contrast to the role of modifications contained in RNA from thermophiles, which is to reduce regional RNA flexibility and provide structural stability to RNA for adaptation to high temperature.
-
Quantitative measurement of Dihydrouridine in RNA using isotope dilution liquid chromatography–mass spectrometry (LC/MS)
1996Co-Authors: Joseph J. Dalluge, Takeshi Hashizume, James A. MccloskeyAbstract:A method has been developed for the microscale determination of 5,6-Dihydrouridine, the most common post-transcriptional modification in bacterial and euka-ryotic tRNA. The method is based on stable isotope dilution liquid chromatography–mass spectrometry (LC/ MS) using [1,3-15N2]Dihydrouridine and [1,3-15N2]uridine as internal standards. RNA samples were enzymatically digested to nucleosides before addition of the internal standards and subsequently analyzed by LC/MS with selected ion monitoring of protonated molecular ions of the labeled and unlabeled nucleosides. Sample quantities of ∼1 pmol tRNA and 5 pmol 23S rRNA were analyzed for mole % Dihydrouridine. Dihydrouridine content of Escherichia coli tRNASerVGA and tRNAThrGGU as controls were measured as 2.03 and 2.84 residues/ tRNA molecule, representing accuracies of 98 and 95%. Overall precision values for the analyses of E.coli tRNASerVGA and E.coli tRNAThrGGU, unfractionated tRNA from E.coli and 23S rRNA from E.coli were within the range 0.43–2.4%. The mole % Dihydrouridine in unfrac-tionated tRNA and 23S rRNA from E.coli were deter-mined as 1.79 and 0.0396%, corresponding to 1.4 and 1.1 residues/RNA molecule respectively
-
Conformational flexibility in RNA: the role of Dihydrouridine
1996Co-Authors: Joseph J. Dalluge, James A. Mccloskey, Takeshi Hashizume, Alan E. Sopchik, Darrell R. DavisAbstract:In order to further understand the structural role of the modified nucleoside Dihydrouridine in RNA the solution conformations of Dp and ApDpA were analyzed by one-and two-dimensional proton NMR spectroscopy and compared with those of the related uridine-containing compounds. The analyses indicate that Dihydrouridine significantly destabilizes the C3′-endo sugar con-formation associated with base stacked, ordered, A-type helical RNA. Equilibrium constants (Keq = [C2′-endo]/[C3′-endo]) for C2′-endo–C3′-endo interconversion at 25C for Dp, the 5′-terminal A of ApDpA and D in ApDpA are 2.08, 1.35 and 10.8 respectively. Stabilization of the C2′-endo form was shown to be enhanced at low temperature, indicating that C2′-endo is the thermo-dynamically favored conformation for Dihydrouridine. ∆H values show that for Dp the C2′-endo sugar conformation is stabilized by 1.5 kcal/mol compared with Up. This effect is amplified for D in the oligonucleotide ApDpA and propagated to the 5′-neighboring A, with stabilization of the C2′-endo form by 5.3 kcal/mol for D and 3.6 kcal/mol for the 5′-terminal A. Post-tran-scriptional formation of Dihydrouridine therefore represents a biological strategy opposite in effect to ribose methylation, 2-thiolation or pseudouridylation, all of which enhance regional stability through stabilization of the C3′-endo conformer. Dihydrouridine effectively promotes the C2′-endo sugar conformation, allowing for greater conformational flexibility and dynamic motion in regions of RNA where tertiary interactions and loop formation must be simultaneously accomodated
Antonio Federico - One of the best experts on this subject based on the ideXlab platform.
-
a novel heteroplasmic trnaleu cun mtdna point mutation associated with chronic progressive external ophthalmoplegia
Biochemical and Biophysical Research Communications, 2005Co-Authors: Elena Cardaioli, Paola Da Pozzo, Elena Radi, Maria Teresa Dotti, Antonio FedericoAbstract:We have sequenced all mitochondrial tRNA genes from a patient with chronic progressive external ophthalmoplegia (CPEO) and mitochondrial myopathy, who had no detectable large mtDNA deletions. Direct sequencing failed to detect previously reported mutations and showed a heteroplasmic mutation at nucleotide 12,276 in the tRNA(Leu(CUN)) gene, in the Dihydrouridine stem, which is highly conserved through the species during evolution. RFLP analyses confirmed that 18% of muscle mtDNA harbored the mutation, while it was absent from DNA of fibroblasts and lymphocytes of the proband and in 110 patients with other encephalomyopathies. To date, besides large and single nucleotide deletions, several point mutations on mitochondrial tRNA genes have been reported in CPEO patients, but only three were in the gene coding for tRNA(Leu(CUN)).
-
A novel heteroplasmic tRNA Leu(CUN) mtDNA point mutation associated with chronic progressive external ophthalmoplegia
'Elsevier BV', 2005Co-Authors: Cardaioli E, Da Pozzo P, Radi E, Antonio FedericoAbstract:We have sequenced all mitochondrial tRNA genes from a patient with chronic progressive external ophthalmoplegia (CPEO) and mitochondrial myopathy, who had no detectable large mtDNA deletions. Direct sequencing failed to detect previously reported mutations and showed a heteroplasmic mutation at nucleotide 12,276 in the tRNA(Leu(CUN)) gene, in the Dihydrouridine stem, which is highly conserved through the species during evolution. RFLP analyses confirmed that 18% of muscle mtDNA harbored the mutation, while it was absent from DNA of fibroblasts and lymphocytes of the proband and in 110 patients with other encephalomyopathies. To date, besides large and single nucleotide deletions, several point mutations on mitochondrial tRNA genes have been reported in CPEO patients, but only three were in the gene coding for tRNA(Leu(CUN))
Theodore J Papenfuss - One of the best experts on this subject based on the ideXlab platform.
-
replication slippage may cause parallel evolution in the secondary structures of mitochondrial transfer rnas
Molecular Biology and Evolution, 1997Co-Authors: J R Macey, Natalia B. Ananjeva, Allan Larson, Theodore J PapenfussAbstract:Presence of the Dihydrouridine (D) stem in the mitochondrial cysteine tRNA is unusually variable among lepidosaurian reptiles. Phylogenetic and comparative analyses of cysteine tRNA gene sequences identify eight parallel losses of the D-stem, resulting in D-arm replacement loops. Sampling within the monophyletic Acrodonta provides no evidence for reversal. Slipped-strand mispairing of noncontiguous repeated sequences during replication or direct replication slippage can explain repeats observed within cysteine tRNAs that contain a D-arm replacement loop. These two mechanisms involving replication slippage can account for the loss of the cysteine tRNA D-stem in several lepidosaurian lineages, and may represent general mechanisms by which the secondary structures of mitochondrial tRNAs are altered.
Mccloskey J A - One of the best experts on this subject based on the ideXlab platform.
-
Quantitative measurement of Dihydrouridine in RNA using isotope dilution liquid chromatography-mass spectrometry (LC/MS).
1996Co-Authors: Dalluge J J, Hashizume T, Mccloskey J AAbstract:A method has been developed for the microscale determination of 5,6-Dihydrouridine, the most common post-transcriptional modification in bacterial and eukaryotic tRNA. The method is based on stable isotope dilution liquid chromatography-mass spectrometry (LC/MS) using [1,3-15N2]Dihydrouridine and [1,3-15N2]uridine as internal standards. RNA samples were enzymatically digested to nucleosides before addition of the internal standards and subsequently analyzed by LC/MS with selected ion monitoring of protonated molecular ions of the labeled and unlabeled nucleosides. Sample quantities of approximately 1 pmol tRNA and 5 pmol 23S rRNA were analyzed for mole% Dihydrouridine. Dihydrouridine content of Escherichia coli tRNASer(VGA) and tRNAThr(GGU) as controls were measured as 2.03 and 2.84 residues/tRNA molecule, representing accuracies of 98 and 95%. Overall precision values for the analyses of E. coli tRNASer(VGA) and E. coli tRNAThr(GGU), unfractionated tRNA from E. coli and 23S rRNA from E. coli were within the range 0.43-2.4%. The mole% Dihydrouridine in unfractionated tRNA and 23S rRNA from E. coli were determined as 1.79 and 0.0396%, corresponding to 1.4 and 1.1 residues/RNA molecule respectively
-
Conformational flexibility in RNA: the role of Dihydrouridine.
1996Co-Authors: Dalluge J J, Hashizume T, Mccloskey J A, Sopchik A E, Davis D RAbstract:In order to further understand the structural role of the modified nucleoside Dihydrouridine in RNA the solution conformations of Dp and ApDpA were analyzed by one- and two-dimensional proton NRM spectroscopy and compared with those of the related uridine-containing compounds. The analyses indicate that Dihydrouridine significantly destabilizes the C3'-endo sugar conformation associated with base stacked, ordered, A-type helical RNA. Equilibrium constants (Keq = [C2'-endo]/[C3'-endo]) for C2'-endo-C3'-endo interconversion at 25 degrees C for Dp, the 5'-terminal A of ApDpA and D in ApDpA are 2.08, 1.35 and 10.8 respectively. Stabilization of the C2'-endo form was shown to be enhanced at low temperature, indicating that C2'-endo is the thermodynamically favored conformation for Dihydrouridine. DeltaH values show that for Dp the C2'-endo sugar conformation is stabilized by 1.5 kcal/mol compared with Up. This effect is amplified for D in the oligonucleotide ApDpA and propagated to the 5'-neighboring A, with stabilization of the C2'-endo form by 5.3 kcal/mol for D and 3.6 kcal/mol for the 5'-terminal A. Post-transcriptional formation of Dihydrouridine therefore represents a biological strategy opposite in effect to ribose methylation, 2-thiolation or pseudouridylation, all of which enhance regional stability through stabilization of the C3'-endo conformer. Dihydrouridine effectively promotes the C2'-endo sugar conformation, allowing for greater conformational flexibility and dynamic motion in regions of RNA where tertiary interactions and loop formation must be simultaneously accommodated