Protein post-translational modifications such as phosphorylation and S-nitrosylation regulate protein functions and cellular programs in eukaryotes.Moreover,extensive evidence suggests crosstalk between these modifica...Protein post-translational modifications such as phosphorylation and S-nitrosylation regulate protein functions and cellular programs in eukaryotes.Moreover,extensive evidence suggests crosstalk between these modifications.However,we lack a comprehensive method for the simultaneous detection and analysis of multiple post-translational modifications.Here,we present an optimized workflow that identifies phosphorylation and S-nitrosylation sites using a novel phosphate affinity tag switch technique.Validation with model proteins and complex biological samples confirmed the high sensitivity,coverage,and reproducibility of this method.Applying this method to Arabidopsis thaliana seedlings revealed 12,552 phosphorylation sites and 6,108 S-nitrosylation sites,representing the largest single-study dataset of S-nitrosylation sites to date.This approach enhances our understanding of post-translational modification dynamics in plant signaling,stress responses,and metabolism.展开更多
基金support of the Science and Technology Program of Guangdong Province(2023A0505090005,2021TQ06N115)the Natural Science Foundation of Guangdong Province(2022A1515110962)+1 种基金the Special Fund for Scientific Innovation Strategy-Construction of High-Level Academy of Agriculture Science(R2021YJ-YB3010,R2023PY-JG025,2023QZ-NK04,GDNKY-ZQQZ-K5)Modern Seed Industry Innovation Capability Enhancement Project of Guangdong Academy of Agricultural Sciences.
文摘Protein post-translational modifications such as phosphorylation and S-nitrosylation regulate protein functions and cellular programs in eukaryotes.Moreover,extensive evidence suggests crosstalk between these modifications.However,we lack a comprehensive method for the simultaneous detection and analysis of multiple post-translational modifications.Here,we present an optimized workflow that identifies phosphorylation and S-nitrosylation sites using a novel phosphate affinity tag switch technique.Validation with model proteins and complex biological samples confirmed the high sensitivity,coverage,and reproducibility of this method.Applying this method to Arabidopsis thaliana seedlings revealed 12,552 phosphorylation sites and 6,108 S-nitrosylation sites,representing the largest single-study dataset of S-nitrosylation sites to date.This approach enhances our understanding of post-translational modification dynamics in plant signaling,stress responses,and metabolism.
