Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins
Abstract Allosteric conformational change is an important paradigm in the regulation of protein function, which is typically triggered by the binding of small cofactors, metal ions or protein partners. Here, we found those conformational transitions can be effectively monitored by 19F NMR, facilitat...
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Nature Portfolio
2024-12-01
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| Series: | Communications Biology |
| Online Access: | https://doi.org/10.1038/s42003-024-07331-x |
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| author | Xichun Liu Pengfei Guo Qiufan Yu Shu-Qin Gao Hong Yuan Xiangshi Tan Ying-Wu Lin |
| author_facet | Xichun Liu Pengfei Guo Qiufan Yu Shu-Qin Gao Hong Yuan Xiangshi Tan Ying-Wu Lin |
| author_sort | Xichun Liu |
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| description | Abstract Allosteric conformational change is an important paradigm in the regulation of protein function, which is typically triggered by the binding of small cofactors, metal ions or protein partners. Here, we found those conformational transitions can be effectively monitored by 19F NMR, facilitated by a site-specific 19F incorporation strategy at the protein C-terminus using asparaginyl endopeptidase (AEP). Three case studies show that C-terminal 19F-nuclei can reveal protein dynamics not only adjacent but also distal to C-terminus, including those occurring in a hemoprotein neuroglobin (Ngb), calmodulin (CaM), and a cobalt metalloregulator (CoaR) responding to both cobalt and tetrapyrrole. In Ngb, the heme orientation disorder is affected by missense mutations that perturb backbone rigidity or surface charges close to the heme axial ligands. In CaM, the C-terminal 19F-nuclei is an ideal probe for detecting the binding states of Ca2+, peptides and inhibitors. Furthermore, multiple 19F-moieties were incorporated into the two domains of CoaR, revealing the intrinsically disordered C-terminal metal binding tail might be an allosteric conformational switch to maintain cobalt homeostasis and balance corrinoid biosynthesis. This study demonstrates that the AEP-based 19F-modification strategy can be applied to various targets to study allosteric regulation, especially for those biological processes modulated by the protein C-terminus. |
| format | Article |
| id | doaj-art-2353aaa895174442a1e7171b044a088f |
| institution | OA Journals |
| issn | 2399-3642 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Nature Portfolio |
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| series | Communications Biology |
| spelling | doaj-art-2353aaa895174442a1e7171b044a088f2025-08-20T02:31:50ZengNature PortfolioCommunications Biology2399-36422024-12-017111210.1038/s42003-024-07331-xSite-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteinsXichun Liu0Pengfei Guo1Qiufan Yu2Shu-Qin Gao3Hong Yuan4Xiangshi Tan5Ying-Wu Lin6School of Chemistry and Chemical Engineering, University of South ChinaSchool of Chemistry and Chemical Engineering, University of South ChinaSchool of Chemistry and Chemical Engineering, University of South ChinaKey Lab of Protein Structure and Function of Universities in Hunan Province, Hengyang Medical School, University of South ChinaDepartment of Chemistry & Institute of Biomedical Science, Fudan UniversityDepartment of Chemistry & Institute of Biomedical Science, Fudan UniversitySchool of Chemistry and Chemical Engineering, University of South ChinaAbstract Allosteric conformational change is an important paradigm in the regulation of protein function, which is typically triggered by the binding of small cofactors, metal ions or protein partners. Here, we found those conformational transitions can be effectively monitored by 19F NMR, facilitated by a site-specific 19F incorporation strategy at the protein C-terminus using asparaginyl endopeptidase (AEP). Three case studies show that C-terminal 19F-nuclei can reveal protein dynamics not only adjacent but also distal to C-terminus, including those occurring in a hemoprotein neuroglobin (Ngb), calmodulin (CaM), and a cobalt metalloregulator (CoaR) responding to both cobalt and tetrapyrrole. In Ngb, the heme orientation disorder is affected by missense mutations that perturb backbone rigidity or surface charges close to the heme axial ligands. In CaM, the C-terminal 19F-nuclei is an ideal probe for detecting the binding states of Ca2+, peptides and inhibitors. Furthermore, multiple 19F-moieties were incorporated into the two domains of CoaR, revealing the intrinsically disordered C-terminal metal binding tail might be an allosteric conformational switch to maintain cobalt homeostasis and balance corrinoid biosynthesis. This study demonstrates that the AEP-based 19F-modification strategy can be applied to various targets to study allosteric regulation, especially for those biological processes modulated by the protein C-terminus.https://doi.org/10.1038/s42003-024-07331-x |
| spellingShingle | Xichun Liu Pengfei Guo Qiufan Yu Shu-Qin Gao Hong Yuan Xiangshi Tan Ying-Wu Lin Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins Communications Biology |
| title | Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins |
| title_full | Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins |
| title_fullStr | Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins |
| title_full_unstemmed | Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins |
| title_short | Site-specific incorporation of 19F-nuclei at protein C-terminus to probe allosteric conformational transitions of metalloproteins |
| title_sort | site specific incorporation of 19f nuclei at protein c terminus to probe allosteric conformational transitions of metalloproteins |
| url | https://doi.org/10.1038/s42003-024-07331-x |
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