A spectroscopic analysis of the interaction between the human regulatory proteins RACK1 and Ki-1/57
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Flávia C. Nery
Abstract
Ki-1/57 is a 57-kDa cytoplasmic and nuclear protein associated with protein kinase activity and is hyper-phosphorylated on Ser/Thr residues upon cellular activation. In previous studies we identified the receptor of activated kinase-1 (RACK1), a signaling adaptor protein that binds activated PKC, as a protein that interacts with Ki-1/57. Here we demonstrate that the far-UV circular dichroism spectrum of the WD repeat-containing RACK1 protein shows an unusual positive ellipticity at 229 nm, which in other proteins of the WD family has been attributed to surface tryptophans that are quenchable by N-bromosuccinimide (NBS). As well as NBS, in vitro binding of 6×His-Ki-1/57(122–413) and 6×His-Ki-1/57(264–413) can also quench the positive ellipticity of the RACK1 spectrum. We generated a model of RACK1 by homology modeling using a G protein β subunit as template. Our model suggests the family-typical seven-bladed β-propeller, with an aromatic cluster around the central tunnel that contains four Trp residues (17, 83, 150, 170), which are likely involved in the interaction with Ki-1/57.
References
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Articles in the same Issue
- Protein aggregation in crowded environments
- Nitrite, a naturally occurring precursor of nitric oxide that acts like a ‘prodrug’
- Functional studies of the small subunit of EcoHK31I DNA methyltransferase
- Functional analysis of amino acid residues at the dimerisation interface of KpnI DNA methyltransferase
- Conformation and stability of the Streptococcus pyogenes pSM19035-encoded site-specific β recombinase, and identification of a folding intermediate
- Tyr-48, a conserved residue in ribotoxins, is involved in the RNA-degrading activity of α-sarcin
- Pathogenicity of catalytic antibodies: catalytic activity of Bence Jones proteins from myeloma patients with renal impairment can elicit cytotoxic effects
- Transgenic expression of gallerimycin, a novel antifungal insect defensin from the greater wax moth Galleria mellonella, confers resistance to pathogenic fungi in tobacco
- Catalytic pathways of Euphorbia characias peroxidase reacting with hydrogen peroxide
- Biochemical and pharmacological characterization of the human bradykinin subtype 2 receptor produced in mammalian cells using the Semliki Forest virus system
- A spectroscopic analysis of the interaction between the human regulatory proteins RACK1 and Ki-1/57
- Subcellular localisation of human inositol 1,4,5-trisphosphate 3-kinase C: species-specific use of alternative export sites for nucleo-cytoplasmic shuttling indicates divergent roles of the catalytic and N-terminal domains
- The gating effect of calmodulin and calcium on the connexin50 hemichannel
- C-Terminal fusion of eGFP to the bradykinin B2 receptor strongly affects down-regulation but not receptor internalization or signaling
- Angiotensin I-converting enzyme inhibitor peptides derived from the endostatin-containing NC1 fragment of human collagen XVIII
- μ-Calpain binds to lipid bilayers via the exposed hydrophobic surface of its Ca2+-activated conformation
- Cathepsin L splice variants in human breast cell lines
