Advanced search
Start date
(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Molecular insights on CALX-CBD12 interdomain dynamics from MD simulations, RDCs, and SAXS

Full text
de Souza Degenhardt, Maximilia F. [1] ; Vitale, Phelipe A. M. [2] ; Abiko, Layara A. [2] ; Zacharias, Martin [3] ; Sattler, Michael [4] ; Oliveira, Cristiano L. P. [1] ; Salinas, Roberto K. [2]
Total Authors: 7
[1] Univ Sao Paulo, Inst Phys, Dept Expt Phys, Sao Paulo - Brazil
[2] Univ Sao Paulo, Inst Chem, Dept Biochem, Sao Paulo - Brazil
[3] Tech Univ Munich, Phys Dept, Munich - Germany
[4] Tech Univ Munich, Chem Dept, Munich - Germany
Total Affiliations: 4
Document type: Journal article
Source: BIOPHYSICAL JOURNAL; v. 120, n. 17, p. 3664-3675, SEP 7 2021.
Web of Science Citations: 0

Na+/Ca2+ exchangers (NCXs) are secondary active transporters that couple the translocation of Nathorn with the transport of Ca2+ in the opposite direction. The exchanger is an essential Ca2+ extrusion mechanism in excitable cells. It consists of a transmembrane domain and a large intracellular loop that contains two Ca2+-binding domains, CBD1 and CBD2. The two CBDs are adjacent to each other and form a two-domain Ca2+ sensor called CBD12. Binding of intracellular Ca2+ to CBD12 activates the NCX but inhibits the NCX of Drosophila, CALX. NMR spectroscopy and SAXS studies showed that CALX and NCX CBD12 constructs display significant interdomain flexibility in the apo state but assume rigid interdomain arrangements in the Ca2+-bound state. However, detailed structure information on CBD12 in the apo state is missing. Structural characterization of proteins formed by two or more domains connected by flexible linkers is notoriously challenging and requires the combination of orthogonal information from multiple sources. As an attempt to characterize the conformational ensemble of CALX-CBD12 in the apo state, we applied molecular dynamics (MD) simulations, NMR (H-1-N-15 residual dipolar couplings), and small-angle x-ray scattering (SAXS) data in a combined strategy to select an ensemble of conformations in agreement with the experimental data. This joint approach demonstrated that CALX-CBD12 preferentially samples closed conformations, whereas the wide-open interdomain arrangement characteristic of the Ca2+-bound state is less frequently sampled. These results are consistent with the view that Ca2+ binding shifts the CBD12 conformational ensemble toward extended conformers, which could be a key step in the NCXs' allosteric regulation mechanism. This strategy, combining MD with NMR and SAXS, provides a powerful approach to select ensembles of conformations that could be applied to other flexible multidomain systems. (AU)

FAPESP's process: 18/16092-5 - Investigating colloidal systems by scattering methods
Grantee:Cristiano Luis Pinto de Oliveira
Support Opportunities: Regular Research Grants
FAPESP's process: 16/24531-3 - Structural and biophysics properties of native and modified lipoproteins
Grantee:Antonio Martins Figueiredo Neto
Support Opportunities: Research Projects - Thematic Grants
FAPESP's process: 19/19968-1 - Structural studies of the Na+/Ca2+ exchanger by high-resolution nuclear magnetic resonance spectroscopy
Grantee:Roberto Kopke Salinas
Support Opportunities: Regular Research Grants
FAPESP's process: 17/05614-8 - Na +/Ca2+ exchangers: relationship between structure and function
Grantee:Phelipe Augusto Mariano Vitale
Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
FAPESP's process: 16/07490-1 - Structural studies of the Na+/Ca2+ exchanger and of proteins from the Zika virus by Nuclear Magnetic Resonance spectroscopy in solution
Grantee:Roberto Kopke Salinas
Support Opportunities: Regular Research Grants