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May 20th, 2025
Version: 2
University of Massachusetts, Amherst
biochemistry
biorxiv

A domain-swapped CaMKII conformation facilitates linker-mediated allosteric regulation

Nguyen, V. C. B.Open in Google Scholar•Özden, C.Open in Google Scholar•Dong, K.Open in Google Scholar•Koc, O.Open in Google Scholar•Torres-Ocampo, A. P.Open in Google Scholar•Dziedzic, N.Open in Google Scholar•Flaherty, D.Open in Google Scholar•Huang, J.Open in Google Scholar•Sankura, S.Open in Google Scholar•Abromson, N. L.Open in Google Scholaret al.

Memory formation, fertilization, and cardiac function rely on precise Ca2+ signaling and subsequent Ca2+/calmodulin-dependent protein kinase II (CaMKII) activation. Ca2+ sensitivity of the four CaMKII paralogs in mammals has been linked to the length of the variable linker region that is a hot spot for alternative splicing. In this study, we determined that the position of charged residues within the linker modulates the Ca2+/CaM sensitivity. We present an X-ray crystal structure of the full-length CaMKII{delta} holoenzyme consisting of domain-swapped dimers within a dodecameric complex. In this structure, the kinase domain of one subunit is docked onto the hub domain of an adjacent subunit, providing an additional interface within the holoenzyme. Mutations at the hub equatorial and lateral interfaces led to alterations in the stoichiometry of CaMKII holoenzyme as well as Ca2+/CaM sensitivity. Using molecular dynamics (MD) to compare domain-swapped to non-domain-swapped CaMKIIs, we demonstrate that the domain-swapped conformation facilitates an interaction between the calmodulin binding region and the linker region. Based on MD simulations and small-angle X-ray scattering (SAXS) measurements, we propose a model where the position of charges on the linker region drives an interaction with the regulatory segment that modulates the degree of autoinhibition. Finally, we use live-cell imaging to show that the activation profiles we observe in vitro are recapitulated in cells. Our findings provide a new framework for understanding allosteric regulation of CaMKII by the linker region in Ca2+-sensitive cells.

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