Molecular dynamics(MD)simulations and anisotropic thermal diffusion dynamics(ATD)simulations were performed on the wild TrpR and its 75 residue mutant(mTrpR)to investigate TrpR longrange effects.The ATD result shows t...Molecular dynamics(MD)simulations and anisotropic thermal diffusion dynamics(ATD)simulations were performed on the wild TrpR and its 75 residue mutant(mTrpR)to investigate TrpR longrange effects.The ATD result shows that the mTrpR has higher fluctuation than the wild TrpR,and its helix chainⅡF has particular disorder.It is obvious that the 75 residue of wild TrpR and mTrpR affects the protein dynamics flexibilities by the long-range effects.The ATD and MD both confirm that the differences in the size of side-chain and three-dimensional structures of two different 75 residues in the wild TrpR and mTrpR will spread to the entire protein by way of the long-range effects.Long-range effect affects the protein side chain interaction,conformational changes,flexibilities and secondary structures.Further,the ATD result also shows that each 75 residue of the symmetric homodimer has the same effect,and the two 75 residues have a positive correlation in long-range regulating processes.The residues 48,50,71,79 in chainⅠof wild TrpR and residues 45,72,80 in chainⅡof mTrpR play important roles in long-range interaction processes.展开更多
In order to investigate the recognition mechanism and the relationship between structure and function of minihelix DNA with Tryptophanyl-tRNA Synthetase (TrpRS), TrpRS from Bacillus Subtilis was purified. Four minihel...In order to investigate the recognition mechanism and the relationship between structure and function of minihelix DNA with Tryptophanyl-tRNA Synthetase (TrpRS), TrpRS from Bacillus Subtilis was purified. Four minihelix DNAs were chemically synthesized and the photoreactive reagent s4T was incorporated into three of them at the positions of G73, T72 and T55 corresponding to tRNATrp. The apparatus for uv crosslinking was devised and the parameters for uv crosslinking were optimized. The results indicated that the G73 and T72 base of minihelix DNA interacted with TrpRS directly. The uv crosslinking reaction was improved by the dose of uv irradiation and the concentration of both TrpRS and minihelix DNA.展开更多
Abstract The details of species- spe- cific aminoacylation in Oryza sativa mitochondrial tRNATrp by bacterial and eukaryotic (cytoplasm) tryptophanyl-tRNA synthetases (TrpRS) were inves- tigated. Seven single or multi...Abstract The details of species- spe- cific aminoacylation in Oryza sativa mitochondrial tRNATrp by bacterial and eukaryotic (cytoplasm) tryptophanyl-tRNA synthetases (TrpRS) were inves- tigated. Seven single or multiple mutations of three bases (G73, U72, A 68) were made in O. sativa mi- tochondrial tRNATrp to the corresponding nucleotides present in human tRNATrp. In vitro transcripts of these mutant genes were tryptophanylated by Bacil- lus subtilis and human tryptophanyl-tRNA syntheta- ses (TrpRS), and the kinetic parameters were deter- mined. The results showed that the aminoacylation of seven mutant transcripts by B. subtilis TrpRS was 53.33%―99.79% less efficient than that by wild-type O. sativa mitochondrial tRNATrp, but was 4―330 times more efficient than that by human TrpRS. The mutant MPH7 (G73, U72 and C68 in O. sativa mito- chondrial tRNA were all replaced by the counterpart residues from human tRNATrp and showed a great change in aminoacylation efficiency. Our results in- dicate that the species-specific identity elements of O. sativa mitochondrial tRNATrp are similar to bacterial and eukaryotic (cytoplasm). They are mainly located at the discriminator base, the first and the fifth pairs of bases, the discriminator base G73, two bases in the acceptor stem G1/U72 and U5/A68. Our results also provide new data in support of the hypothesis that mitochondrial tRNATrp is of eubacterial origin.展开更多
基金supported by the Innovation project of Henan Agricultural University(No.30600982)PhD Start-up Foundation of Henan Agricultural University(30600780)2023 Instrument Operator Capability lmprovement Project(SYS2023T04)
文摘Molecular dynamics(MD)simulations and anisotropic thermal diffusion dynamics(ATD)simulations were performed on the wild TrpR and its 75 residue mutant(mTrpR)to investigate TrpR longrange effects.The ATD result shows that the mTrpR has higher fluctuation than the wild TrpR,and its helix chainⅡF has particular disorder.It is obvious that the 75 residue of wild TrpR and mTrpR affects the protein dynamics flexibilities by the long-range effects.The ATD and MD both confirm that the differences in the size of side-chain and three-dimensional structures of two different 75 residues in the wild TrpR and mTrpR will spread to the entire protein by way of the long-range effects.Long-range effect affects the protein side chain interaction,conformational changes,flexibilities and secondary structures.Further,the ATD result also shows that each 75 residue of the symmetric homodimer has the same effect,and the two 75 residues have a positive correlation in long-range regulating processes.The residues 48,50,71,79 in chainⅠof wild TrpR and residues 45,72,80 in chainⅡof mTrpR play important roles in long-range interaction processes.
基金This work was supported by the National Natural Science Foundation of China (Grant No. 39730120) the Chinese Academy of Sciences (Grant No. KSCX 2-2-04).
文摘In order to investigate the recognition mechanism and the relationship between structure and function of minihelix DNA with Tryptophanyl-tRNA Synthetase (TrpRS), TrpRS from Bacillus Subtilis was purified. Four minihelix DNAs were chemically synthesized and the photoreactive reagent s4T was incorporated into three of them at the positions of G73, T72 and T55 corresponding to tRNATrp. The apparatus for uv crosslinking was devised and the parameters for uv crosslinking were optimized. The results indicated that the G73 and T72 base of minihelix DNA interacted with TrpRS directly. The uv crosslinking reaction was improved by the dose of uv irradiation and the concentration of both TrpRS and minihelix DNA.
基金supported by the Natural Science Foundation of Zhejiang Province(Grant No.302103).
文摘Abstract The details of species- spe- cific aminoacylation in Oryza sativa mitochondrial tRNATrp by bacterial and eukaryotic (cytoplasm) tryptophanyl-tRNA synthetases (TrpRS) were inves- tigated. Seven single or multiple mutations of three bases (G73, U72, A 68) were made in O. sativa mi- tochondrial tRNATrp to the corresponding nucleotides present in human tRNATrp. In vitro transcripts of these mutant genes were tryptophanylated by Bacil- lus subtilis and human tryptophanyl-tRNA syntheta- ses (TrpRS), and the kinetic parameters were deter- mined. The results showed that the aminoacylation of seven mutant transcripts by B. subtilis TrpRS was 53.33%―99.79% less efficient than that by wild-type O. sativa mitochondrial tRNATrp, but was 4―330 times more efficient than that by human TrpRS. The mutant MPH7 (G73, U72 and C68 in O. sativa mito- chondrial tRNA were all replaced by the counterpart residues from human tRNATrp and showed a great change in aminoacylation efficiency. Our results in- dicate that the species-specific identity elements of O. sativa mitochondrial tRNATrp are similar to bacterial and eukaryotic (cytoplasm). They are mainly located at the discriminator base, the first and the fifth pairs of bases, the discriminator base G73, two bases in the acceptor stem G1/U72 and U5/A68. Our results also provide new data in support of the hypothesis that mitochondrial tRNATrp is of eubacterial origin.
