Two TPX2-Dependent Switches Control the Activity of Aurora A

Xue, Xu; Wang, Xia; Xiao, Zhengtao; Li, Yan; Wang, Yonghua

doi:10.1371/journal.pone.0016757

Cited by 18 publications

(22 citation statements)

References 29 publications

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“…S5A and S6B). Residues 141-143 in the glycine-rich loop are highly flexible and Lys-143 has been shown to be a switch in the case of ATP binding in Aurora A (19). We also analyzed the second most favorable docked pose in which the felodipine was hydrogen bonded to the residues His-201 and Trp-128.…”

Section: Validation Of Sers Results Through Molecular Docking and MDmentioning

confidence: 99%

SERS and MD simulation studies of a kinase inhibitor demonstrate the emergence of a potential drug discovery tool

Dhanasekaran

Siddhanta

Kishore

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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We demonstrate the use of surface-enhanced Raman spectroscopy (SERS) as an excellent tool for identifying the binding site of small molecules on a therapeutically important protein.As an example, we show the specific binding of the common antihypertension drug felodipine to the oncogenic Aurora A kinase protein via hydrogen bonding interactions with Tyr-212 residue to specifically inhibit its activity. Based on SERS studies, molecular docking, molecular dynamics simulation, biochemical assays, and point mutation-based validation, we demonstrate the surface-binding mode of this molecule in two similar hydrophobic pockets in the Aurora A kinase. These binding pockets comprise the same unique hydrophobic patches that may aid in distinguishing human Aurora A versus human Aurora B kinase in vivo. The application of SERS to identify the specific interactions between small molecules and therapeutically important proteins by differentiating competitive and noncompetitive inhibition demonstrates its ability as a complementary technique. We also present felodipine as a specific inhibitor for oncogenic Aurora A kinase. Felodipine retards the rate of tumor progression in a xenografted nude mice model. This study reveals a potential surface pocket that may be useful for developing small molecules by selectively targeting the Aurora family kinases.vibrational spectroscopy | structure-activity relationship | ligand binding U nderstanding the mechanism of ligand binding to proteins is imperative for designing new molecules or screening potential drug molecules from available databases. We have used surface-enhanced Raman spectroscopy (SERS), which is a highly sensitive technique, to understand the binding of the commonly used hypertension drug, felodipine, to Aurora A kinase. Although NMR (1), X-ray crystallography (2), surface plasmon resonance (3), and fluorescence (4) are experimental techniques used to explore protein-drug interactions and each of these techniques provides unique information about the proteinligand interaction, a common problem of these techniques is the requirement of a high-protein concentration or the incorporation of secondary tagged molecules and a protein size limit. SERS has been traditionally used for the ultrasensitive detection of analytes. However, it can also be used to examine the protein-small molecule interactions and elucidate the mechanism (5-7). A commonly debated aspect is that SERS does not provide complete vibrational information compared with resonant Raman or normal Raman spectroscopy. Despite the limited information from SERS, which can be performed in proteins at extremely low concentrations in their active state, the competitive binding versus noncompetitive binding and specific changes in protein upon ligand binding can be explained. This approach is extremely effective when combined with molecular dynamics (MD) simulations and the structural information of the protein. The usefulness of this combination is that drugs can be screened for therapeutic applications. This paper provid...

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Section: Validation Of Sers Results Through Molecular Docking and MDmentioning

confidence: 99%

SERS and MD simulation studies of a kinase inhibitor demonstrate the emergence of a potential drug discovery tool

Dhanasekaran

Siddhanta

Kishore

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…These include centrosome multiplication1011 and maturation12131415, bipolar spindle formation161718, stability and function192021. Indeed, while interphase cells contain a significant cytoplasmic pool of AURKA2223, the kinase localises to distinct structures during cell cycle progression242526.…”

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confidence: 99%

Allosteric modulation of AURKA kinase activity by a small-molecule inhibitor of its protein-protein interaction with TPX2

Janecek

Rossmann

Sharma

et al. 2016

Sci Rep

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The essential mitotic kinase Aurora A (AURKA) is controlled during cell cycle progression via two distinct mechanisms. Following activation loop autophosphorylation early in mitosis when it localizes to centrosomes, AURKA is allosterically activated on the mitotic spindle via binding to the microtubule-associated protein, TPX2. Here, we report the discovery of AurkinA, a novel chemical inhibitor of the AURKA-TPX2 interaction, which acts via an unexpected structural mechanism to inhibit AURKA activity and mitotic localization. In crystal structures, AurkinA binds to a hydrophobic pocket (the ‘Y pocket’) that normally accommodates a conserved Tyr-Ser-Tyr motif from TPX2, blocking the AURKA-TPX2 interaction. AurkinA binding to the Y- pocket induces structural changes in AURKA that inhibit catalytic activity in vitro and in cells, without affecting ATP binding to the active site, defining a novel mechanism of allosteric inhibition. Consistent with this mechanism, cells exposed to AurkinA mislocalise AURKA from mitotic spindle microtubules. Thus, our findings provide fresh insight into the catalytic mechanism of AURKA, and identify a key structural feature as the target for a new class of dual-mode AURKA inhibitors, with implications for the chemical biology and selective therapeutic targeting of structurally related kinases.

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“…TPX2 is shown as a beige ribbon, bound to the N-terminal domain far from the active site. In this position the activating loop is protected from dephosphorylation by negative Aurora-A regulators, and is available for interaction with Aurora-A substrates [37–43]. The order of TPX2 binding and Aurora-A phosphorylation in vivo is not known, though it has been proposed that low levels of auto-phosphorylation of Aurora-A occur at centrosomes in the early stages of mitosis, followed by allosteric activation by TPX2 promoting high levels of actions as the centrosome assembles the spindle microtubules [32].…”

Section: Mitotic Activation Of Aurora-a: the Important Role Of T28mentioning

confidence: 99%

Issues in interpreting thein vivoactivity of Aurora-A

Shagisultanova

Dunbrack

Golemis

2014

Expert Opinion on Therapeutic Targets

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Introduction Based on its role as a mitotic regulatory kinase, overexpressed and associated with aneuploidy in cancer, small molecule inhibitors have been developed for Aurora-A (AURKA) kinase. In preclinical and clinical assessments, these agents have shown efficacy in inducing stable disease or therapeutic response. In optimizing the use of Aurora-A inhibitors, it is critical to have robust capacity to measure the kinase activity of Aurora-A in tumors. Areas covered we provide an overview of molecular mechanisms of mitotic and non-mitotic activation of Aurora-A kinase, and interaction of Aurora-A with its regulatory partners. Typically, Aurora-A activity is measured by use of phospho-antibodies targeting an auto-phosphorylated T288 epitope. However, recent studies have identified alternative means of Aurora-A activation control, including allosteric regulation by partners, phosphorylation on alternative activating residues (S51, S98), dephosphorylation on inhibitory sites (S342), and T288 phosphorylation by alternative kinases such as Pak enzymes. Additional work has shown that the relative abundance of Aurora-A partners can affect the activity of Aurora-A inhibitors, and that Aurora-A activation also occurs in interphase cells. Expert opinion Taken together, this work suggests the need for comprehensive analysis of Aurora-A activity and expression of Aurora-A partners in order to stratify patients for likely therapeutic response.

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Two TPX2-Dependent Switches Control the Activity of Aurora A

Cited by 18 publications

References 29 publications

SERS and MD simulation studies of a kinase inhibitor demonstrate the emergence of a potential drug discovery tool

SERS and MD simulation studies of a kinase inhibitor demonstrate the emergence of a potential drug discovery tool

Allosteric modulation of AURKA kinase activity by a small-molecule inhibitor of its protein-protein interaction with TPX2

Issues in interpreting thein vivoactivity of Aurora-A

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