Characterization of CaMKIIα holoenzyme stability

Torres‐Ocampo, Ana P.; Ozden, C.; Hommer, Alexandra; Gardella, Anne; Lapinskas, Emily; Samkutty, Alfred; Esposito, Edward A.; Garman, S.C.; Stratton, Margaret M.

doi:10.1002/pro.3869

Cited by 27 publications

(32 citation statements)

References 42 publications

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“…1B). This hinge is more rigid (0.29 + 0.002 nm) than the Lat-Dim hinge (0.355 + 0.016 nm) consistent with previous work 9,12 or the AD-KD "clip" contact (0.292 + 0.005 nm). I used principal component analysis (PCA) to classify long-range collective motions within the tetramer.…”

Section: Resultssupporting

confidence: 89%

Conformational Spread Drives the Evolution of the Calcium-calmodulin Protein Kinase II

Khan

2021

Preprint

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The calcium calmodulin (Ca2+CAM) dependent protein kinase II (CaMKII) decodes Ca2+ frequency oscillations. It has a central role in learning. I matched residue and organismal evolution to collective motions deduced from the atomic structure of the human CaMKIIa holoenzyme. Protein dynamic simulations and bioinformatic analysis showed its stacked ring architecture conformationally couples kinase domains (KDs) via its central hub. The simulations revealed underlying b-sheet collective motions in the hub ab association domain (AD) map onto a coevolved residue network and partition it into two distinct sectors. The holoenzyme evolved in metazoans by stabilization of ancient enzyme dimers and fold elongation to create a second, metastable sector for ring assembly. Continued evolution targeted the ring contacts for lateral conformational spread. The a isoform, predominantly expressed in the brain, emerged last and evolved rapidly in sync with the poikilotherm-homeotherm jump in the evolution of memory. The correlation between CaMKII dynamics and phylogenetics argues single residue evolution fine-tunes hub conformational spread. The central role of CaMKII ringed architecture In the brain could be to increase Ca2+ frequency response range for complex learning functions.

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Section: Resultssupporting

confidence: 89%

Conformational Spread Drives the Evolution of the Calcium-calmodulin Protein Kinase II

Khan

2021

Preprint

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“…This indicates that these hydrophobic residues shift to accommodate substrate binding, which is likely a large contribution to an observed gain in kinase domain stability upon GluN2B binding (Fig. S2e), similar to what we have previously observed with regulatory segment binding, which is mediated by L290 in the regulatory segment [36]. All effectors studied here have a leucine at the -5 position.…”

Section: Discussionsupporting

confidence: 87%

CaMKII binds both substrates and activators at the active site

Ozden

Sloutsky

Santos

et al. 2020

Preprint

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Ca2+/calmodulin dependent protein kinase II (CaMKII) is a signaling protein that is required for successful long-term memory formation. Ca2+/CaM activates CaMKII by binding to its regulatory segment, thereby making the substrate binding pocket available. One exceptional feature of this kinase is that two binding partners have been shown persistently activate CaMKII after the Ca2+ stimulus dissipates. The molecular details of this phenomenon are unclear. Despite having a large variety of interaction partners, the specificity of CaMKII has not been structurally well-characterized. To this end, we solved X-ray crystal structures of the CaMKII kinase domain bound to four different binding partners: a peptide substrate, a substrate/activator, a substrate/binding partner, and an inhibitor (AMPA-type glutamate receptor, NMDA-type glutamate receptor, Tiam1, and Densin-180). We show that all four binding partners use similar interactions to bind across the substrate binding pocket of the CaMKII active site. We generated a sequence alignment based on our structural observations, which revealed conserved interactions across these binding partners. The structures presented here shed much-needed light on the interaction between CaMKII and its binding partners. These observations will be crucial in guiding further biological experiments.

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“…Consistent with this, a construct consisting of just the hub and the regulatory segment was shown to undergo spontaneous subunit exchange ( Bhattacharyya et al, 2016 ). Recent differential scanning calorimetry and mass photometry experiments have shown that while the isolated hub is very stable, the holoenzyme, especially with the linker present, is unstable and undergoes dissociation ( Torres-Ocampo et al, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

Breakage of the oligomeric CaMKII hub by the regulatory segment of the kinase

Karandur

Bhattacharyya

Xia

et al. 2020

eLife

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Ca2+/calmodulin dependent protein kinase II (CaMKII) is an oligomeric enzyme with crucial roles in neuronal signaling and cardiac function. Previously, we showed that activation of CaMKII triggers the exchange of subunits between holoenzymes, potentially increasing the spread of the active state (Stratton et al. 2014; Bhattacharyya et al. 2016). Using mass spectrometry, we show now that unphosphorylated and phosphorylated peptides derived from the CaMKII-α regulatory segment bind to the CaMKII-α hub and break it into smaller oligomers. Molecular dynamics simulations show that the regulatory segments dock spontaneously at the interface between hub subunits, trapping large fluctuations in hub structure. Single-molecule fluorescence intensity analysis of CaMKII-α expressed in mammalian cells shows that activation of CaMKII-α results in the destabilization of the holoenzyme. Our results suggest that release of the regulatory segment by activation and phosphorylation allows it to destabilize the hub, producing smaller assemblies that might reassemble to form new holoenzymes.

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Characterization of CaMKIIα holoenzyme stability

Cited by 27 publications

References 42 publications

Conformational Spread Drives the Evolution of the Calcium-calmodulin Protein Kinase II

Conformational Spread Drives the Evolution of the Calcium-calmodulin Protein Kinase II

CaMKII binds both substrates and activators at the active site

Breakage of the oligomeric CaMKII hub by the regulatory segment of the kinase

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