Regulation of cell fate requires the establishment and erasure of 5-methylcytosine(5mC) in genomic DNA.The formation of 5mC is achieved by DNA cytosine methyltransferases(DNMTs),whereas the removal of5mC can be accomp...Regulation of cell fate requires the establishment and erasure of 5-methylcytosine(5mC) in genomic DNA.The formation of 5mC is achieved by DNA cytosine methyltransferases(DNMTs),whereas the removal of5mC can be accomplished by various pathways.Aside from ten-eleven translocation(TET)-mediated oxidation of 5mC followed by thymine DNA glycosylase(TDG)-initiated base excision repair(BER),the direct deformylation of 5-formylcytosine(5fC) and decarboxylation of 5-carboxylcytosine(5caC) have also been discovered as the novel DNA demethylation pathways.Although these novel demethylation pathways have been identified in stem cells and somatic cells,their precise roles in regulating cell fate remain unclear.Here,we differentiate mouse embryonic stem cells(mESCs) into mouse embryoid bodies(mEBs),followed by further differentiation into mouse neural stem cells(mNSCs) and finally into mouse neurons(mNeurons).During this sequential differentiation process,we employ probe molecules,namely2'-fluorinated 5-formylcytidine(F-5fC) and 2'-fluorinated 5-carboxyldeoxycytidine(F-5caC),for metabolic labeling.The results of mass spectrometry(MS) analysis demonstrate the deformylation and decarboxylation activities are progressively decreased and increased respectively during differentiation process,and this opposite demethylation tendency is not associated with DNMTs and TETs.展开更多
The fatty alk（a/e）ne biosynthesis pathway found in cyanobacteria gained tremendous attention in recent years as a promising alternative approach for biofuel production. Cyanobacterial aldehyde-deformylating oxygena...The fatty alk（a/e）ne biosynthesis pathway found in cyanobacteria gained tremendous attention in recent years as a promising alternative approach for biofuel production. Cyanobacterial aldehyde-deformylating oxygenase （cADO）, which catalyzes the conversion of Cn fatty aldehyde to its corresponding Cn-1 alk（ale）ne, is a key enzyme in that pathway. Due to its low activity, alk（a/e）ne production by cADO is an inefficient process. Previous biochemical and structural investi. gations of cADO have provided some information on its catalytic reaction. However, the details of its cata- lytic processes remain unclear. Here we report five crystal structures of cADO from the Synechococcus elongates strain PCC7942 in both its iron-free and iron-bound forms, representing different states during its catalytic process. Structural comparisons and functional enzyme assays indicate that Giu144, one of the iron-coordinating residues, plays a vital role in the catalytic reaction of cADO. Moreover, the helix where Glu144 resides exhibits two distinct conformations that correlates with the different binding states of the di-iron center in cADO structures. Therefore, our results provide a structural explanation for the highly labile feature of cADO di-iron center, which we pro- posed to be related to its low enzymatic activity. On the basis of our structural and biochemical data, a possible catalytic process of cADO was proposed, which could aid the design of cADO with improved activity.展开更多
Manganese(Ⅲ)-peroxo complexes are invoked as key intermediates in the enzymatic cycles of Mncontaining enzymes, and the synthesis of reactive manganese(Ⅲ)-peroxo complexes with rationally designed ligand has been of...Manganese(Ⅲ)-peroxo complexes are invoked as key intermediates in the enzymatic cycles of Mncontaining enzymes, and the synthesis of reactive manganese(Ⅲ)-peroxo complexes with rationally designed ligand has been of great interest in the communities of bioinorganic and biomimetic chemistry.Herein, we designed a novel pentadentate aminobenzimidazole ligand and obtained its manganese(Ⅱ)complex, which was successfully applied in the synthesis of a reactive manganese(Ⅲ)-peroxo complex by treatment with hydrogen peroxide in the presence of triethylamine. The manganese(Ⅲ)-peroxo complex was well characterized with various spectroscopic techniques, including ultraviolet-visible(UV-vis)spectrophotometry, coldspray ionization time-of-flight mass spectrometry(CSI-TOF MS), and continuous wave electron paramagnetic resonance(CW-EPR) spectroscopy. Besides, its reactivity in aldehyde deformylation was investigated, demonstrating second-order kinetics in the reaction with 2-phenylpropionaldehyde and affording acetophenone as the sole product.展开更多
A peroxonickel(III)complex bearing the Me6-trien ligand,an open-chain analogue of the macrocyclic 12-TMC ligand,was synthesized and characterized by several physical methods,such as UV-vis,resonance Raman,ESI-MS and E...A peroxonickel(III)complex bearing the Me6-trien ligand,an open-chain analogue of the macrocyclic 12-TMC ligand,was synthesized and characterized by several physical methods,such as UV-vis,resonance Raman,ESI-MS and EPR.Density functional theory calculations provide geometric and electronic configurations of peroxonickel(III)complexes.The reactivity of the peroxonickel(III)intermediate was examined in oxidative nucleophilic reactions(e.g.,aldehyde deformylation).By comparing the reactivities of the open-chain and macrocyclic peroxonickel(III)complexes under identical reaction conditions,we were able to demonstrate that the open-chain peroxonickel(III)complex is much more reactive than the macrocyclic analogue in aldehyde deformylation.This result provides intriguing aspects of the reactivity differences for open-chain vs.macrocyclic systems of metal complexes.展开更多
We report a substrate-dependent annulation system where 6-substituents of 2H-1,4-benzoxazines dictate divergent pathways with ynamides.Non-methoxy substrates undergo TBSOTf/Zn(OTf)_(2)-catalyzed[2+2]annulation/ring ex...We report a substrate-dependent annulation system where 6-substituents of 2H-1,4-benzoxazines dictate divergent pathways with ynamides.Non-methoxy substrates undergo TBSOTf/Zn(OTf)_(2)-catalyzed[2+2]annulation/ring expansion to form 2H-1,6-benzoxazocines,while 6-methoxy derivatives preferentially yield 4-aminoquinolines via a TBSOTf-catalyzed[4+2]annulation/deformylation pathway.This electronic effect-driven selectivity operates under mild conditions with high fidelity.The method provides orthogonal access to two medicinally important heterocycle classes from identical precursors,features broad functional group tolerance,demonstrates scalability(up to 1 mmol scale),and eliminates the need for transition metals.The selectivity may originate from the differential stabilization of intermediates by the 6-substituent.展开更多
基金supported by the National Key R&D Program of China (Nos.2022YFC3400700,2022YFA0806600)the National Natural Science Foundation of China (No.22074110)+3 种基金Guangdong Basic and Applied Basic Research Foundation (No.2022A1515110550)Central Public-interest Scientific Institution Basal Research Fund,South China Sea Fisheries Research Institute,CAFS (No.2021TS02)Guangzhou Basic and Applied Basic Research Foundation (No.2023A04J1337)Central Public-interest Scientific Institution Basal Research Fund,CAFS (No.2023TD78)。
文摘Regulation of cell fate requires the establishment and erasure of 5-methylcytosine(5mC) in genomic DNA.The formation of 5mC is achieved by DNA cytosine methyltransferases(DNMTs),whereas the removal of5mC can be accomplished by various pathways.Aside from ten-eleven translocation(TET)-mediated oxidation of 5mC followed by thymine DNA glycosylase(TDG)-initiated base excision repair(BER),the direct deformylation of 5-formylcytosine(5fC) and decarboxylation of 5-carboxylcytosine(5caC) have also been discovered as the novel DNA demethylation pathways.Although these novel demethylation pathways have been identified in stem cells and somatic cells,their precise roles in regulating cell fate remain unclear.Here,we differentiate mouse embryonic stem cells(mESCs) into mouse embryoid bodies(mEBs),followed by further differentiation into mouse neural stem cells(mNSCs) and finally into mouse neurons(mNeurons).During this sequential differentiation process,we employ probe molecules,namely2'-fluorinated 5-formylcytidine(F-5fC) and 2'-fluorinated 5-carboxyldeoxycytidine(F-5caC),for metabolic labeling.The results of mass spectrometry(MS) analysis demonstrate the deformylation and decarboxylation activities are progressively decreased and increased respectively during differentiation process,and this opposite demethylation tendency is not associated with DNMTs and TETs.
基金We would like to thank Yi Han and Shengquan Liu at the Institute of Biophysics, CAS and the staffs at Shanghai Synchrotron Radiation Facility and Photo Factory, Japan, respectively, for help during X-ray data collection. We also thank Wei Shao at Beijing Center for Physical and Chemical Analysis for help with the GC analysis, Zhen Xue at Institute of Botany, CAS for providing help for the GC-QqQ- MS/MS analysis, and Hongzhi Zhang at Institute of Geographic Sciences and Natural Resources Research, CAS for technical support with the metal content analysis by ICP-OES. This work was supported by the National Basic Research Program （973 Program） （Nos. 2011CBA00902 and 2011CBA00907）, Strategic Priority Research Program of the Chinese Academy of Sciences （XDB08020302）, National Natural Science Foundation of China（Grant Nos. 31021062 and 31170765）, and CAS Cross and Coop- eration Team for Scientific Innovation （Y31102110A）.
文摘The fatty alk（a/e）ne biosynthesis pathway found in cyanobacteria gained tremendous attention in recent years as a promising alternative approach for biofuel production. Cyanobacterial aldehyde-deformylating oxygenase （cADO）, which catalyzes the conversion of Cn fatty aldehyde to its corresponding Cn-1 alk（ale）ne, is a key enzyme in that pathway. Due to its low activity, alk（a/e）ne production by cADO is an inefficient process. Previous biochemical and structural investi. gations of cADO have provided some information on its catalytic reaction. However, the details of its cata- lytic processes remain unclear. Here we report five crystal structures of cADO from the Synechococcus elongates strain PCC7942 in both its iron-free and iron-bound forms, representing different states during its catalytic process. Structural comparisons and functional enzyme assays indicate that Giu144, one of the iron-coordinating residues, plays a vital role in the catalytic reaction of cADO. Moreover, the helix where Glu144 resides exhibits two distinct conformations that correlates with the different binding states of the di-iron center in cADO structures. Therefore, our results provide a structural explanation for the highly labile feature of cADO di-iron center, which we pro- posed to be related to its low enzymatic activity. On the basis of our structural and biochemical data, a possible catalytic process of cADO was proposed, which could aid the design of cADO with improved activity.
基金financially supported by the National Natural Science Foundation of China(No.21473226)Natural Science Foundation of Jiangsu Province(No.BK20161261).We acknowledge Prof
文摘Manganese(Ⅲ)-peroxo complexes are invoked as key intermediates in the enzymatic cycles of Mncontaining enzymes, and the synthesis of reactive manganese(Ⅲ)-peroxo complexes with rationally designed ligand has been of great interest in the communities of bioinorganic and biomimetic chemistry.Herein, we designed a novel pentadentate aminobenzimidazole ligand and obtained its manganese(Ⅱ)complex, which was successfully applied in the synthesis of a reactive manganese(Ⅲ)-peroxo complex by treatment with hydrogen peroxide in the presence of triethylamine. The manganese(Ⅲ)-peroxo complex was well characterized with various spectroscopic techniques, including ultraviolet-visible(UV-vis)spectrophotometry, coldspray ionization time-of-flight mass spectrometry(CSI-TOF MS), and continuous wave electron paramagnetic resonance(CW-EPR) spectroscopy. Besides, its reactivity in aldehyde deformylation was investigated, demonstrating second-order kinetics in the reaction with 2-phenylpropionaldehyde and affording acetophenone as the sole product.
基金financial support from the NRF(2017R1A2B4005441 and 2018R1A5A1025511)the Ministry of Science,ICT and Future Planning(CGRC 2016M3D3A01913243)of Korea.T.O.acknowledges the support of“Strategic Young Researcher Overseas Visits Program for Accelerating Brain Circulation”.
文摘A peroxonickel(III)complex bearing the Me6-trien ligand,an open-chain analogue of the macrocyclic 12-TMC ligand,was synthesized and characterized by several physical methods,such as UV-vis,resonance Raman,ESI-MS and EPR.Density functional theory calculations provide geometric and electronic configurations of peroxonickel(III)complexes.The reactivity of the peroxonickel(III)intermediate was examined in oxidative nucleophilic reactions(e.g.,aldehyde deformylation).By comparing the reactivities of the open-chain and macrocyclic peroxonickel(III)complexes under identical reaction conditions,we were able to demonstrate that the open-chain peroxonickel(III)complex is much more reactive than the macrocyclic analogue in aldehyde deformylation.This result provides intriguing aspects of the reactivity differences for open-chain vs.macrocyclic systems of metal complexes.
基金the Natural Science Foundation of Henan Province(No.252300420229)for financial support.
文摘We report a substrate-dependent annulation system where 6-substituents of 2H-1,4-benzoxazines dictate divergent pathways with ynamides.Non-methoxy substrates undergo TBSOTf/Zn(OTf)_(2)-catalyzed[2+2]annulation/ring expansion to form 2H-1,6-benzoxazocines,while 6-methoxy derivatives preferentially yield 4-aminoquinolines via a TBSOTf-catalyzed[4+2]annulation/deformylation pathway.This electronic effect-driven selectivity operates under mild conditions with high fidelity.The method provides orthogonal access to two medicinally important heterocycle classes from identical precursors,features broad functional group tolerance,demonstrates scalability(up to 1 mmol scale),and eliminates the need for transition metals.The selectivity may originate from the differential stabilization of intermediates by the 6-substituent.
