0576929 - FGÚ 2024 RIV US eng J - Článek v odborném periodiku
Petrvalská, Olivia - Honzejková, K. - Koupilová, N. - Herman, P. - Obšilová, Veronika - Obšil, Tomáš
14-3-3 protein inhibits CaMKK1 by blocking the kinase active site with its last two C-terminal helices.
Protein Science. Roč. 32, č. 11 (2023), č. článku e4805. ISSN 0961-8368. E-ISSN 1469-896X
Grant CEP: GA ČR(CZ) GA19-00121S
Výzkumná infrastruktura: CIISB II - 90127
Institucionální podpora: RVO:67985823
Klíčová slova: 14-3-3 proteins * calcium/calmodulin-dependent protein kinase * CaMKK * fluorescence spectroscopy * hydrogen/deuterium exchange coupled to MS * protein–protein interaction * SAXS
Obor OECD: Biochemistry and molecular biology
Impakt faktor: 4.5, rok: 2023
Způsob publikování: Open access
https://doi.org/10.1002/pro.4805

Ca2+/CaM-dependent protein kinase kinases 1 and 2 (CaMKK1 and CaMKK2) phosphorylate and enhance the catalytic activity of downstream kinases CaMKI, CaMKIV, and protein kinase B. Accordingly, CaMKK1 and CaMKK2 regulate key physiological and pathological processes, such as tumorigenesis, neuronal morphogenesis, synaptic plasticity, transcription factor activation, and cellular energy homeostasis, and promote cell survival. Both CaMKKs are partly inhibited by phosphorylation, which in turn triggers adaptor and scaffolding protein 14-3-3 binding. However, 14-3-3 binding only significantly affects CaMKK1 function. CaMKK2 activity remains almost unchanged after complex formation for reasons still unclear. Here, we aim at structurally characterizing CaMKK1:14-3-3 and CaMKK2:14-3-3 complexes by SAXS, H/D exchange coupled to MS, and fluorescence spectroscopy. The results revealed that complex formation suppresses the interaction of both phosphorylated CaMKKs with Ca2+/CaM and affects the structure of their kinase domains and autoinhibitory segments. But these effects are much stronger on CaMKK1 than on CaMKK2 because the CaMKK1:14-3-3γ complex has a more compact and rigid structure in which the active site of the kinase domain directly interacts with the last two C-terminal helices of the 14-3-3γ protein, thereby inhibiting CaMKK1. In contrast, the CaMKK2:14-3-3 complex has a looser and more flexible structure, so 14-3-3 binding only negligibly affects the catalytic activity of CaMKK2. Therefore, Ca2+/CaM binding suppression and the interaction of the kinase active site of CaMKK1 with the last two C-terminal helices of 14-3-3γ protein provide the structural basis for 14-3-3-mediated CaMKK1 inhibition.
Trvalý link: https://hdl.handle.net/11104/0346310