With rapid developments of emerging technologies like synthetic biology,the demand for DNA polymerases with superior activities including higher thermostability and processivity has increased significantly.Thus,ration...With rapid developments of emerging technologies like synthetic biology,the demand for DNA polymerases with superior activities including higher thermostability and processivity has increased significantly.Thus,rational optimization of the performance of DNA polymerase is of great interest.Nuclear magnetic resonance(NMR)spectroscopy is a powerful technique used for studying protein structure and dynamics.It provides the atomic resolution information of enzymes under their functional solution environment to reveal the active sites(hot spots)of the enzyme,which could be further used for optimizing the performance of enzymes.In our previous work,we identified hot spot residues of Pyrococcus furiosus DNA polymerase(Pfu pol).We aim to employ these binding hot spots to screen for co-factors of Pfu pol,particularly targeting those molecules exhibiting weak intermolecular interactions.To validate this concept,we first demonstrated the feasibility of utilizing hot spot residues as screening probes for auxiliary factors by employing the well-characterized Tween-20 as a model system.Employing these hot spots as probes,two new co-factors,the heat shock protein TkHSP20 from Thermococcus Kodakaraensis and the chemical chaperone L-arginine,are identified to interact with Pfu pol to boost its performance in amplifying long DNA fragments by enhancing the thermal stability and the processivity of the Pfu pol.This NMR-based approach requires no prior assignment information of target enzymes,guiding the rational exploration of novel cofactors for Pfu pol.Moreover,our approach is not dependent on structural data or bioinformatics.Therefore,it has significant potential for application in various enzymes to expedite the progress in enzyme engineering.展开更多
在全球价值链重构与数字技术革命双重驱动下,我国产业转型升级已进入以协同创新驱动高质量发展阶段。从产业协同集聚新视角出发,选取2010—2023年我国31个省级面板数据,实证检验产业协同集聚对我国对外直接投资(Outward Foreign Direct ...在全球价值链重构与数字技术革命双重驱动下,我国产业转型升级已进入以协同创新驱动高质量发展阶段。从产业协同集聚新视角出发,选取2010—2023年我国31个省级面板数据,实证检验产业协同集聚对我国对外直接投资(Outward Foreign Direct Investment,OFDI)的影响。结果表明,生产性服务业与制造业协同集聚对我国OFDI具有显著促进作用。机制分析表明,产业协同集聚通过推动技术创新与全要素生产率提升影响OFDI,数字经济水平会强化协同集聚对OFDI的影响;异质性分析表明,我国东部地区与胡焕庸线东侧地区的协同集聚显著正向促进OFDI;金融业,交通运输、仓储和邮电业,批发,信息传输、软件和信息技术服务业与制造业协同集聚对OFDI具有显著推动效果。展开更多
基金supported by the Strategic Priority Research Program of the Chinese Academy of Sciences XDB0540000Natural Science Foundation of China grants 22327901,22174151 and 21991080Hubei Provincial Natural Science Foundation of China 2023AFA041。
文摘With rapid developments of emerging technologies like synthetic biology,the demand for DNA polymerases with superior activities including higher thermostability and processivity has increased significantly.Thus,rational optimization of the performance of DNA polymerase is of great interest.Nuclear magnetic resonance(NMR)spectroscopy is a powerful technique used for studying protein structure and dynamics.It provides the atomic resolution information of enzymes under their functional solution environment to reveal the active sites(hot spots)of the enzyme,which could be further used for optimizing the performance of enzymes.In our previous work,we identified hot spot residues of Pyrococcus furiosus DNA polymerase(Pfu pol).We aim to employ these binding hot spots to screen for co-factors of Pfu pol,particularly targeting those molecules exhibiting weak intermolecular interactions.To validate this concept,we first demonstrated the feasibility of utilizing hot spot residues as screening probes for auxiliary factors by employing the well-characterized Tween-20 as a model system.Employing these hot spots as probes,two new co-factors,the heat shock protein TkHSP20 from Thermococcus Kodakaraensis and the chemical chaperone L-arginine,are identified to interact with Pfu pol to boost its performance in amplifying long DNA fragments by enhancing the thermal stability and the processivity of the Pfu pol.This NMR-based approach requires no prior assignment information of target enzymes,guiding the rational exploration of novel cofactors for Pfu pol.Moreover,our approach is not dependent on structural data or bioinformatics.Therefore,it has significant potential for application in various enzymes to expedite the progress in enzyme engineering.
文摘在全球价值链重构与数字技术革命双重驱动下,我国产业转型升级已进入以协同创新驱动高质量发展阶段。从产业协同集聚新视角出发,选取2010—2023年我国31个省级面板数据,实证检验产业协同集聚对我国对外直接投资(Outward Foreign Direct Investment,OFDI)的影响。结果表明,生产性服务业与制造业协同集聚对我国OFDI具有显著促进作用。机制分析表明,产业协同集聚通过推动技术创新与全要素生产率提升影响OFDI,数字经济水平会强化协同集聚对OFDI的影响;异质性分析表明,我国东部地区与胡焕庸线东侧地区的协同集聚显著正向促进OFDI;金融业,交通运输、仓储和邮电业,批发,信息传输、软件和信息技术服务业与制造业协同集聚对OFDI具有显著推动效果。
