Characterization of acidic ribosomal proteins from three developmental stages of the medfly Ceratitis capitata

Abstract

Acidic proteins extracted with 0.5 M NH4Cl and 50% ethanol from ribosomes of the medfly Ceratitis capitata showed two major bands of MW 15 and 17 kD after SDS electrophoresis. Isoelectrofocusing of acidic proteins resolved two groups of bands at pH 4.5 and 3.5. Similar patterns were observed both from the acidic ribosomal protein fraction and from total ribosomes, treated with RNase. Treatment with alkaline phosphatase reduced the number of bands with a shift to a higher pI, indicating dephosphorylation. The phosphorylation pattern of the acidic proteins changed at three different stages of development, six day larvae, white pupae and 0‐2 days old embryos. The two protein groups correspond to multi‐phosphorylated forms of eucaryotic acidic ribosomal proteins P1 and P2. This was shown by immunoblotting with specific monoclonal antibodies.

Molecular cloning of the ribosomal P-proteins MgP1, MgP2, MgP0, and superoxide dismutase (SOD) in the mussel Mytilus galloprovincialis and analysis of MgP0 at stress conditions

“…Total ribosomes were isolated from gills of the mussel M. galloprovincialis as previously described (Kouyanou et al, 1998;. Briefly, animal tissue (gills, 10 g) was homogenized in liquid nitrogen and suspended in 2 ml/g of tissue, in 100 mM Tris/HCl pH 7.4, 100 mM KCl, 12.5 mM MgCl 2 , 5 mM β-mercaptoethanol (buffer 1), containing protease inhibitors (pepstatin 1.25 mg/ml, leupeptin 1.25 mg/ml, aprotinin 23 TIU/ml) and 0.5 μΜ PMSF in DMSO.…”

“…Phosphorylation seems to have a regulatory role also in Zea mays, with a possible impact on the control of developmental processes: acidic protein P1/P2 phosphorylation is the main step that regulates their function in axes during germination (Montoya-García et al, 2002;Szick-Miranda and BaileySerres, 2001). P1 and P2 proteins are also phosphorylated in the insects Ceratitis capitata and Bombyx mori (Koumarianou, et al, 2007;Kouyanou, et al, 1998), while they are conservative enough to restore viability in conditional mutant strains of S. cerevisiae, substituting the endogenous proteins (Gagou et al, 2000;Koumarianou et al, 2007, Kouyanou et al, 2003.…”

Cloning and characterization of the ribosomal protein CcP0 of the medfly Ceratitis capitata

“…Ribosomes from 6‐day‐old larvae of the medfly C. capitata were prepared and analysed by isoelectrofocusing in 5% acrylamide gels, in the presence of ampholines 2%, pH 3.0–10.0 (Pharmacia), as described by Kouyanou et al . (1998).…”

“…Ribosomes from 6-day-old larvae of the medfly C. capitata were prepared and analysed by isoelectrofocusing in 5% acrylamide gels, in the presence of ampholines 2%, pH 3.0 -10.0 (Pharmacia), as described by Kouyanou et al (1998). The estimation of the protein bands' pIs was performed by measuring the pH of aqueous solutions, in which the respective isoelectrofocusing gel bands had been dipped and agitated continuously for half an hour at R / T. Ribosomes and total E. coli lysates were also resolved by SDS-PAGE in 15% acrylamide gels.…”

“…4). The purified ribosomes showed two additional, not well resolved bands of 17 kDa and 15 kDa, corresponding to the acidic CcP1 and CcP2 proteins, respectively, of the medfly (Kouyanou et al ., 1998).…”