The expression changes of Rars gene in ischemia-injured neurons were investigated by detecting its translational product arginyl-t RNA synthetase(Arg RS), and the inhibitory effects of ischemic preconditioning(IPC...The expression changes of Rars gene in ischemia-injured neurons were investigated by detecting its translational product arginyl-t RNA synthetase(Arg RS), and the inhibitory effects of ischemic preconditioning(IPC) on Rars gene were explored. Both IPC model and prolonged ischemia(PI) model were established by using the classic oxygen glucose deprivation(OGD) method. The primary cultured neurons were assigned into the following groups: the experimental group(IPC+PI group), undergoing PI after a short period of IPC; the conditional control group(PI control group), subjected to PI without IPC; blank control group, the normally cultured neurons. The Rars transcriptional activities and Arg RS expression levels were measured at different time points after re-oxygenation(3 h/6 h/12 h/24 h). Data were collected and statistically analyzed. Compared to the blank control group, the Rars activities and Arg RS levels were significantly increased in PI control group, peaking at the time point of 6 h after re-oxygenation. Rars activities and Arg RS levels were significantly lower in the experimental group than in the PI control group at different time points after re-oxygenation. PI insult can induce an escalating activity of Rars and lead to Arg RS over-expression in primary cultured neurons. IPC can inhibit the increased Rars activity and down-regulate Arg RS expression of ischemia-insulted neurons. This mechanism may confer ischemic tolerance on neurons.展开更多
Aminoacyl-tRNA syntheses (AARS) can catalyze the adenosine triphosphate (ATP)-dependent acylation of their cognate tRNA(s) with a specific amino acid. They can be seen as an index to reflect the energy metabolic...Aminoacyl-tRNA syntheses (AARS) can catalyze the adenosine triphosphate (ATP)-dependent acylation of their cognate tRNA(s) with a specific amino acid. They can be seen as an index to reflect the energy metabolic rate of ischemic brain cells in ischemic penumbra. This study ex- amined the relationship between arginyl-tRNA synthetase (ArgRS), one of the AARS, and cerebral ischemia in rats. The model of middle cerebral artery occlusion (MCAO) was established in rats. The expression levels of ArgRS protein and mRNA were detected in rat brain tissues at different time points following MCAO by Western blotting and RT-PCR, respectively. The results showed that the MCAO model was successfully established. Western blotting and RT-PCR analysis revealed that the ArgRS protein and mRNA were expressed in brain cells in both ischemic and normal penumbra tissues. The expression levels of ArgRS protein and mRNA peaked at 6 h after MCAO and decreased gradually. At 24 h, the expression levels of ArgRs protein and mRNA in ischemic penumbral tissues were lower than those in normal tissues. The expression levels of ArgRS mRNA and protein in ischemic penumbra var- ied with ischemic time, suggesting that the energy metabolism of brain cells in penumbra changed dy- namically after ischemia to ensure the endogenous self-protection of the body. The brain oxygen supply should be improved as soon as possible, especially within 6-12 h after ischemia, so as to meet the de- mand for energy metabolism in ischemic penumbra and make sure the cell structure remains stable.展开更多
The arginyl-tRNA synthetase (ArgRS) catalyzes the esterification reaction between L-arginine and its cog- nate tRNAArg. Previously reported structures of ArgRS shed considerable light on the tRNA recognition mech- a...The arginyl-tRNA synthetase (ArgRS) catalyzes the esterification reaction between L-arginine and its cog- nate tRNAArg. Previously reported structures of ArgRS shed considerable light on the tRNA recognition mech- anism, while the aspect of amino acid binding in ArgRS remains largely unexplored. Here we report the first crystal structure of E. coli ArgRS (eArgRS) complexed with L-arginine, and a series of mutational studies using isothermal titration calorimetry (ITC). Combined with previously reported work on ArgRS, our results eluci- dated the structural and functional roles of a series of important residues in the active site, which furthered our understanding of this unique enzyme.展开更多
The covalent modification of E. coli arginyl-tRNA synthetase by the 2',3'-dialdehydederivative of tRNA^(Arg) (tRNA_(ox)^(Arg)) resulted in the complete inactivation of the ATP-PPi ex-change and aminoacylation ...The covalent modification of E. coli arginyl-tRNA synthetase by the 2',3'-dialdehydederivative of tRNA^(Arg) (tRNA_(ox)^(Arg)) resulted in the complete inactivation of the ATP-PPi ex-change and aminoacylation activities of the enzyme. Sodium dodecyl sulfate polyacrylamide gelelectrophoresis of the ArgRS-tRNA_(ox)^(Arg) covalent complexes indicated that two bands simulta-neously appeared on the gel parallel with inactivation corresponding to different higher mo-lecular weights. This result was different from that of the other aminoacyl-tRNA synthetaselabeling systems as previously reported. Upon the ribonuclease treatment of the modifiedArgRS, less than 15% of both the initial ATP-PPi exchange and aminocylation activities wererecovered. During the whole process of labeling and RNase treatment, the two activities ofthe enzyme were closely associated.展开更多
基金supported by the National Natural Science Foundation of China(No.81371453)
文摘The expression changes of Rars gene in ischemia-injured neurons were investigated by detecting its translational product arginyl-t RNA synthetase(Arg RS), and the inhibitory effects of ischemic preconditioning(IPC) on Rars gene were explored. Both IPC model and prolonged ischemia(PI) model were established by using the classic oxygen glucose deprivation(OGD) method. The primary cultured neurons were assigned into the following groups: the experimental group(IPC+PI group), undergoing PI after a short period of IPC; the conditional control group(PI control group), subjected to PI without IPC; blank control group, the normally cultured neurons. The Rars transcriptional activities and Arg RS expression levels were measured at different time points after re-oxygenation(3 h/6 h/12 h/24 h). Data were collected and statistically analyzed. Compared to the blank control group, the Rars activities and Arg RS levels were significantly increased in PI control group, peaking at the time point of 6 h after re-oxygenation. Rars activities and Arg RS levels were significantly lower in the experimental group than in the PI control group at different time points after re-oxygenation. PI insult can induce an escalating activity of Rars and lead to Arg RS over-expression in primary cultured neurons. IPC can inhibit the increased Rars activity and down-regulate Arg RS expression of ischemia-insulted neurons. This mechanism may confer ischemic tolerance on neurons.
基金supported by the Natural Science Foundation of China(No.81371453)
文摘Aminoacyl-tRNA syntheses (AARS) can catalyze the adenosine triphosphate (ATP)-dependent acylation of their cognate tRNA(s) with a specific amino acid. They can be seen as an index to reflect the energy metabolic rate of ischemic brain cells in ischemic penumbra. This study ex- amined the relationship between arginyl-tRNA synthetase (ArgRS), one of the AARS, and cerebral ischemia in rats. The model of middle cerebral artery occlusion (MCAO) was established in rats. The expression levels of ArgRS protein and mRNA were detected in rat brain tissues at different time points following MCAO by Western blotting and RT-PCR, respectively. The results showed that the MCAO model was successfully established. Western blotting and RT-PCR analysis revealed that the ArgRS protein and mRNA were expressed in brain cells in both ischemic and normal penumbra tissues. The expression levels of ArgRS protein and mRNA peaked at 6 h after MCAO and decreased gradually. At 24 h, the expression levels of ArgRs protein and mRNA in ischemic penumbral tissues were lower than those in normal tissues. The expression levels of ArgRS mRNA and protein in ischemic penumbra var- ied with ischemic time, suggesting that the energy metabolism of brain cells in penumbra changed dy- namically after ischemia to ensure the endogenous self-protection of the body. The brain oxygen supply should be improved as soon as possible, especially within 6-12 h after ischemia, so as to meet the de- mand for energy metabolism in ischemic penumbra and make sure the cell structure remains stable.
基金This project was funded by the National Basic Research Program (973 Program) (No. 2010CB912301), the Chinese Academy of Sciences (KSCX2-EW-G-7-2) and the National Natural Science Foundation of China (Grant No. 91219202).
文摘The arginyl-tRNA synthetase (ArgRS) catalyzes the esterification reaction between L-arginine and its cog- nate tRNAArg. Previously reported structures of ArgRS shed considerable light on the tRNA recognition mech- anism, while the aspect of amino acid binding in ArgRS remains largely unexplored. Here we report the first crystal structure of E. coli ArgRS (eArgRS) complexed with L-arginine, and a series of mutational studies using isothermal titration calorimetry (ITC). Combined with previously reported work on ArgRS, our results eluci- dated the structural and functional roles of a series of important residues in the active site, which furthered our understanding of this unique enzyme.
基金Project supported by the National Natural Science Foundation of China.
文摘The covalent modification of E. coli arginyl-tRNA synthetase by the 2',3'-dialdehydederivative of tRNA^(Arg) (tRNA_(ox)^(Arg)) resulted in the complete inactivation of the ATP-PPi ex-change and aminoacylation activities of the enzyme. Sodium dodecyl sulfate polyacrylamide gelelectrophoresis of the ArgRS-tRNA_(ox)^(Arg) covalent complexes indicated that two bands simulta-neously appeared on the gel parallel with inactivation corresponding to different higher mo-lecular weights. This result was different from that of the other aminoacyl-tRNA synthetaselabeling systems as previously reported. Upon the ribonuclease treatment of the modifiedArgRS, less than 15% of both the initial ATP-PPi exchange and aminocylation activities wererecovered. During the whole process of labeling and RNase treatment, the two activities ofthe enzyme were closely associated.