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Article Dans Une Revue BMC Structural Biology Année : 2010

Solution structure of the human signaling protein RACK1

Résumé

Background The adaptor protein RACK1 (receptor of activated kinase 1) was originally identified as an anchoring protein for protein kinase C. RACK1 is a 36 kDa protein, and is composed of seven WD repeats which mediate its protein-protein interactions. RACK1 is ubiquitously expressed and has been implicated in diverse cellular processes involving: protein translation regulation, neuropathological processes, cellular stress, and tissue development. Results In this study we performed a biophysical analysis of human RACK1 with the aim of obtaining low resolution structural information. Small angle X-ray scattering (SAXS) experiments demonstrated that human RACK1 is globular and monomeric in solution and its low resolution structure is strikingly similar to that of an homology model previously calculated by us and to the crystallographic structure of RACK1 isoform A from Arabidopsis thaliana . Both sedimentation velocity and sedimentation equilibrium analytical ultracentrifugation techniques showed that RACK1 is predominantly a monomer of around 37 kDa in solution, but also presents small amounts of oligomeric species. Moreover, hydrodynamic data suggested that RACK1 has a slightly asymmetric shape. The interaction of RACK1 and Ki-1/57 was tested by sedimentation equilibrium. The results suggested that the association between RACK1 and Ki-1/57(122-413) follows a stoichiometry of 1:1. The binding constant (KB) observed for RACK1-Ki-1/57(122-413) interaction was of around (1.5 ± 0.2) × 10 6 M -1 and resulted in a dissociation constant (KD) of (0.7 ± 0.1) × 10 -6 M. Moreover, the fluorescence data also suggests that the interaction may occur in a cooperative fashion. Conclusion Our SAXS and analytical ultracentrifugation experiments indicated that RACK1 is predominantly a monomer in solution. RACK1 and Ki-1/57(122-413) interact strongly under the tested conditions.
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Dates et versions

hal-03751735 , version 1 (17-08-2022)

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Kaliandra Gonçalves, Julio Borges, Julio Cesar Da Silva, Priscila Papa, Gustavo Bressan, et al.. Solution structure of the human signaling protein RACK1. BMC Structural Biology, 2010, 10 (1), pp.15. ⟨10.1186/1472-6807-10-15⟩. ⟨hal-03751735⟩
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