title: Direct observation of Ca(2+) -induced calmodulin conformational transitions in intact Xenopus laevis oocytes by (19) F NMR spectroscopy.
abstract: The Ca(2+) -mediated conformational transition of the protein calmodulin (CaM) is essential to a variety of signal transduction pathways. Whether the transition in living cells is similar to that observed in buffer is not known. Here, we report the direct observation by (19) F NMR spectroscopy of the transition of the Ca(2+) -free and -bound forms in Xenopus laevis oocytes at different Ca(2+) levels. We find that the Ca(2+) -bound CaM population increased greatly upon binding the target protein myosin light-chain kinase (MLCK) at the same Ca(2+) level. Paramagnetic NMR spectroscopy was also exploited for the first time to obtain long-range structural constraints in cells. Our study shows that (19) F NMR spectroscopy can be used to obtain long-range structural constraints in living eukaryotic cells and paves the way for quantification of protein binding constants.
authors.affiliation: Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, National Center for Magnetic Resonance in Wuhan, Wuhan Institute of Physics and Mathematics, Chinese Academy of Sciences, Wuhan 430071 (China); Graduate University of Chinese Academy of Sciences, Beijing 100049 (China).
authors.forename: Yansheng
authors.initials: Y
authors.lastname: Ye
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Liu
authors.forename: Guohua
authors.initials: G
authors.lastname: Xu
authors.forename: Maili
authors.initials: M
authors.lastname: Liu
authors.forename: Conggang
authors.initials: C
authors.lastname: Li
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism*; D005461:Fluorine; D007554:Isotopes; D008958:Models, Molecular; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D009865:Oocytes / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D014982:Xenopus laevis
chemical_list: D002147:Calmodulin; D007554:Isotopes; D005461:Fluorine; D009219:Myosin-Light-Chain Kinase; D002118:Calcium
keywords: NMR spectroscopy; calmodulin; fluorine; protein conformation; signal transduction