More modifiable sites on the nucleoside motif may need to be explored for developing novel(p)ppGpp molecular tools.Herein,we report for the first time that the C7-substituted deazapurine nucleoside triphosphates beari...More modifiable sites on the nucleoside motif may need to be explored for developing novel(p)ppGpp molecular tools.Herein,we report for the first time that the C7-substituted deazapurine nucleoside triphosphates bearing small modifications as substrates could be effectively accepted by RelSeqNTD protein to react with ATP to give pppGpp derivatives with 65%—89%yields.Further structural derivatization via metal-coupling reaction was performed to produce C7-substituted GTP derivatives with larger bulkiness,and those GTP derivatives were also proven to be good substrates of RelSeqNTD protein.Alkynyl modified pppGpp could be coupled with probes by click reactions as the potential molecular tools for fishing proteins in biological research.We further explored whether the C7-alkynyl-pppGpp(pppGEpp)could be recognized by pppGpp interaction proteins.A micromolar level binding affinity(with a KD value of less than 10μM)between pppGpp(pppGEpp)and its binding proteins was obtained from the Isothermal Titration Curve(ITC).All those illustrate that these easily accessible functionalized C7-substituted pppGpp derivatives were suitable tools for further exploring the molecular interaction between pppGpp and its binding proteins.展开更多
基金supported by the National Natural ScienceFoundation of China(U23A20106,22111530224,22304044 and 22473040)the Outstanding Youth Innovation Research Group in Henan Province(242300421005)Doctoral Initiation Fund(202103084).
文摘More modifiable sites on the nucleoside motif may need to be explored for developing novel(p)ppGpp molecular tools.Herein,we report for the first time that the C7-substituted deazapurine nucleoside triphosphates bearing small modifications as substrates could be effectively accepted by RelSeqNTD protein to react with ATP to give pppGpp derivatives with 65%—89%yields.Further structural derivatization via metal-coupling reaction was performed to produce C7-substituted GTP derivatives with larger bulkiness,and those GTP derivatives were also proven to be good substrates of RelSeqNTD protein.Alkynyl modified pppGpp could be coupled with probes by click reactions as the potential molecular tools for fishing proteins in biological research.We further explored whether the C7-alkynyl-pppGpp(pppGEpp)could be recognized by pppGpp interaction proteins.A micromolar level binding affinity(with a KD value of less than 10μM)between pppGpp(pppGEpp)and its binding proteins was obtained from the Isothermal Titration Curve(ITC).All those illustrate that these easily accessible functionalized C7-substituted pppGpp derivatives were suitable tools for further exploring the molecular interaction between pppGpp and its binding proteins.
