A water-mediated allosteric network governs activation of Aurora kinase A

Cyphers, Soreen; Ruff, Emily F.; Behr, Julie M.; Chodera, John D.; Levinson, Nicholas M.

doi:10.1038/nchembio.2296

Cited by 58 publications

(95 citation statements)

References 61 publications

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“…IR spectra of labeled phosphorylated AurA bound to ADP showed predominantly a single absorbance band centered at 2158 cm −1 (Figure 1a). We previously assigned this peak in spectra of unphosphorylated AurA to the DFG-Out form of the kinase, in which the nitrile probe is buried in a hydrophobic pocket (Figure 1b) 21 . Addition of saturating amounts of Tpx2 peptide (residues 1-43 of human Tpx2) to the IR samples caused a dramatic change in the spectra in which the central peak at 2158 cm −1 largely disappears, and two new peaks appear at 2149 cm −1 and 2164 cm −1 (Figure 1a).…”

Section: Resultsmentioning

confidence: 97%

“…We set out to explain how phosphorylation of AurA on T288 leads to a ~100-fold increase in catalytic activity (Supplementary Figure S1) 19,20 . We previously used an infrared (IR) probe that tracks the DFG motif of AurA to show that Tpx2 binding triggers a conformational change from the DFG-Out to the DFG-In state 21 . In this method, a cysteine residue is introduced at position Q185 at the back of the active site of AurA, and chemical labeling is used to introduce a nitrile infrared probe at this position 25 .…”

Section: Resultsmentioning

confidence: 99%

“…Extensive in vitro studies have shown that Tpx2 and phosphorylation act independently to increase AurA kinase activity by up to several hundred-fold 19,20 . Binding of Tpx2 to unphosphorylated AurA triggers a population shift from a DFG-Out to the DFG-In state in solution 21 . Crystal structures of phosphorylated AurA bound to Tpx2 show that the T288 phosphothreonine residue forms extensive ionic interactions unique to the DFG-In state, suggesting that both factors simply stabilize the same active conformation 22–24 .…”

Section: Introductionmentioning

confidence: 99%

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A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation

Ruff

Muretta

Thompson

et al. 2017

Preprint

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23Many eukaryotic protein kinases are activated by phosphorylation on a specific conserved 24 residue in the regulatory activation loop, a post-translational modification thought to stabilize the 25 active DFG-In state of the catalytic domain. Here we use a battery of spectroscopic methods 26. CC-BY 4.0 International license peer-reviewed) is the author/funder. It is made available under aThe copyright holder for this preprint (which was not . http://dx.doi.org/10.1101/205260 doi: bioRxiv preprint first posted online 2 that track different catalytic elements of the kinase domain to show that the ~100-fold activation 27 of the mitotic kinase Aurora A (AurA) by phosphorylation occurs without a population shift to the 28 DFG-In state, and that the activation loop of the activated kinase remains highly dynamic.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation

Ruff

Muretta

Thompson

et al. 2017

Preprint

Self Cite

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“…However, only in the case of TPX2 has the mechanism of activation, and how this synergizes with phosphorylation of T288, been resolved (Bayliss et al , ; Eyers et al , ; Dodson & Bayliss, ). Amino acids (aa) 1–43 of TPX2 stimulate Aurora‐A activity, whether T288 is phosphorylated or not, and TPX2‐binding and T288 phosphorylation together stabilize the αC‐helix and activation loop in a fully ordered conformation (Fig A, left) similar to that observed in active Ser/Thr kinases, such as PKA (Bayliss et al , ; Goldsmith et al , ; Dodson & Bayliss, ; Zorba et al , ; Cyphers et al , ). This mechanism is antagonized by a single domain antibody that traps Aurora‐A into an inactive conformation (Burgess et al , ).…”

Section: Introductionmentioning

confidence: 64%

Mitotic spindle association of TACC3 requires Aurora‐A‐dependent stabilization of a cryptic α‐helix

et al. 2018

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Aurora‐A regulates the recruitment of TACC3 to the mitotic spindle through a phospho‐dependent interaction with clathrin heavy chain (CHC). Here, we describe the structural basis of these interactions, mediated by three motifs in a disordered region of TACC3. A hydrophobic docking motif binds to a previously uncharacterized pocket on Aurora‐A that is blocked in most kinases. Abrogation of the docking motif causes a delay in late mitosis, consistent with the cellular distribution of Aurora‐A complexes. Phosphorylation of Ser558 engages a conformational switch in a second motif from a disordered state, needed to bind the kinase active site, into a helical conformation. The helix extends into a third, adjacent motif that is recognized by a helical‐repeat region of CHC, not a recognized phospho‐reader domain. This potentially widespread mechanism of phospho‐recognition provides greater flexibility to tune the molecular details of the interaction than canonical recognition motifs that are dominated by phosphate binding.

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“…Our results add to data arguing that while the DFG motif is an important determinant of ATP binding its mobilization follows different strategies during activation of EPKs. In AurA, activation by the effector TpX2 involves an equilibrium population shift from the OUT to IN state (46); but activation by phosphorylation triggers a switch between auto-inhibited and activated IN states (44). Multiple strategies for kinase auto-inhibition have also been identified, for example, in the ZAP-70 tyrosine kinase (47).…”

Section: Discussionmentioning

confidence: 99%

Conformational coupling by trans-phosphorylation in calcium calmodulin dependent kinase II

Pandini

Schulman

Khan

2019

Preprint

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31The calcium calmodulin dependent protein kinase II (CaMKII) is a dodecameric holoenzyme 32 important for encoding memory. Its activation, triggered by binding of calcium calmodulin, persists 33 autonomously after calmodulin dissociation. One (receiver) kinase captures and subsequently phos-34 phorylates the regulatory domain peptide of a donor kinase forming a chained dimer as a first stage 35 of autonomous activation. Protein dynamics simulations examined the conformational changes trig-36 gered by dimer formation and phosphorylation, aimed to provide a molecular rationale for human 37 mutations that result in learning disabilities. Ensembles generated from X-ray crystal structures were 38 characterized by network centrality and community analysis. Mutual information related collective 39 motions to local fragment dynamics encoded with a structural alphabet. Implicit solvent tCONCOORD 40 conformational ensembles revealed the dynamic architecture of Inactive kinase domains was co-opted 41 in the activated dimer but the network hub shifted from the nucleotide binding cleft to the captured 42 peptide. Explicit solvent molecular dynamics (MD) showed nucleotide and substrate binding determi-43 nants formed coupled nodes in long-range signal relays between regulatory peptides in the dimer. 44Strain in the extended captured peptide was balanced by reduced flexibility of the receiver kinase C-45 lobe core. The relays were organized around a hydrophobic patch between the captured peptide and 46a key binding helix. The human mutations aligned along the relays. Thus, these mutations could disrupt 47 Author Summary 58Protein kinases play central roles in intracellular signalling. Auto-phosphorylation by bound 59 nucleotide typically precedes phosphate transfer to multiple substrates. Protein conformational 60 changes are central to kinase function, altering binding affinities to change cellular location and shunt 61 from one signal pathway to another. In the brain, the multi-subunit kinase, CaMKII is activated by cal-62 cium calmodulin upon calcium jumps produced by synaptic stimulation. Auto-transphosphorylation of 63 a regulatory peptide enables the kinase to remain activated and mediate long-term behavioural effects 64 after return to basal calcium levels. A database of mutated residues responsible for these effects is 65 difficult to reconcile solely with impaired nucleotide or substrate binding. Therefore, we have compu-66 tationally generated interaction networks to map the conformational plasticity of the kinase domains 67where most mutations localize. The network generated from the atomic structure of a phosphorylated 68 dimer resolves protein sectors based on their collective motions. The sectors link nucleotide and sub-69 strate binding sites in self-reinforcing relays between regulatory peptides. The self-reinforcement is 70 strengthened by phosphorylation consistent with the reported positive cooperativity of kinase activity 71 with calcium-calmodulin concentration. The network gives a better match with the mu...

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A water-mediated allosteric network governs activation of Aurora kinase A

Cited by 58 publications

References 61 publications

A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation

A dynamic mechanism for allosteric activation of Aurora kinase A by activation loop phosphorylation

Mitotic spindle association of TACC3 requires Aurora‐A‐dependent stabilization of a cryptic α‐helix

Conformational coupling by trans-phosphorylation in calcium calmodulin dependent kinase II

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