the acidic ribosomal phosphoproteins from Saccharomyces cerevisiae

Zínker, Samuel

doi:10.1016/0005-2787(80)90099-4

Cited by 46 publications

(47 citation statements)

References 20 publications

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“…Although most ribosomal proteins are found exclusively in ribosomes or in the nucleolus, cytoplasmic pools of Aproteins (or P-proteins) free of ribosomes have been observed in E. coli, S. cerevisiae, A. salina, and HeLa cells (10,40,46,57,63). This suggests that, in contrast to most ribosomal proteins, the acidic ribosomal proteins may be able to exchange on and off of the ribosome in vivo.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, there have been reports that the nonribosomal P-proteins are less phosphorylated than those on the ribosome (46,63) and that phosphorylation seems to increase the activities of P-proteins (32) or their affinities for the ribosome (46). P-proteins are phosphorylated by casein kinase type II, a cyclic AMP-independent kinase which can utilize either ATP or GTP (19,20,22,26,27,35).…”

Section: Discussionmentioning

confidence: 99%

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Human acidic ribosomal phosphoproteins P0, P1, and P2: analysis of cDNA clones, in vitro synthesis, and assembly.

Rich¹,

Steitz²

1987

Mol. Cell. Biol.

273

153

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cDNA clones encoding three antigenically related human ribosomal phosphoproteins (P-proteins) P0, P1, and P2 were isolated and sequenced. P1 and P2 are analogous to Escherichia coli ribosomal protein L7/L12, and P0 is likely to be an analog of LIO. The three proteins have a nearly identical carboxy-terminal 17-amino-acid sequence (KEESEESD(D/E)DMGFGLFD-COOH) that is the basis of their immunological cross-reactivity. The identities of the P1 and P2 cDNAs were confirmed by the strong similarities of their encoded amino acid sequences to published primary structures of the homologous rat, brine shrimp, and Saccharomyces cerevisiae proteins. The P0 cDNA was initially identified by translation of hybrid-selected mRNA and immunoprecipitation of the products. To demonstrate that the coding sequences are full length, the PO, P1, and P2 cDNAs were transcribed in vitro by bacteriophage T7 RNA polymerase and the resulting mRNAs were translated in vitro. The synthetic P0, P1, and P2 proteins were serologically and electrophoretically identical to P-proteins extracted from HeLa cells. These synthetic P-proteins were incorporated into 60S but not 40S ribosomes and also assembled into a complex similar to that described for E. coli L7/L12 and L10.Acidic ribosomal proteins from diverse species have been studied extensively by a number of different techniques. These proteins, called A-proteins (acidic) or P-proteins (the phosphorylated eucaryotic A-proteins), are generally present in multiple copies on the ribosome and have isoelectric points in the range of pH 3 to 5, in contrast to most ribosomal proteins, which are single copy and basic. Aproteins have hydrophobic amino acid compositions (notably about 20% alanine) (34). Because of their distinct chemical properties, A-proteins can be easily dissociated from the ribosome by treatment with high salt and 50% ethanol; the A-proteins remain soluble whereas the remainder of the ribosomes or subunits precipitate (17). This procedure provides a uniform, simple protocol for the identification and isolation of A-proteins from many sources.The best characterized A-protein is the L7/L12 protein of Escherichia coli (L7 is the N-acetylated form of L12 [molecular weight, 12,164]), which has been localized to the stalk of the large (50S) ribosomal subunit (5,38). L7/L12 interacts with several translation factors, some of which bind and hydrolyze GTP, including initiation factor IF2, elongation factors EF-Tu and EF-G, and release factors RF1 and RF2 (5, 59). L7/L12 is required for binding of these factors as well as aminoacyl-tRNA to the ribosome (5, 59). Hydrodynamic and circular dichroism studies have suggested that L7/L12 and other A-proteins are prolate and have high alpha-helical contents (16). However, the crystal structure of the Cterminal domain of L7/L12 has been determined (28), and it is remarkably globular in light of the elongated shape of the whole molecule. The results of intraribosomal proteinprotein cross-linking studies and reconstitution experiments have depicted a complex...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Human acidic ribosomal phosphoproteins P0, P1, and P2: analysis of cDNA clones, in vitro synthesis, and assembly.

Rich¹,

Steitz²

1987

Mol. Cell. Biol.

273

153

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“…Second, acidic proteins of the 60s subunit play an important role in the formation of active ribosomes. It has been observed not only in the yeast cells, but also in other eucaryotic organisms [23,24,471. The specific assembly of the active ribosomal structure from the 'core' particles and 'split proteins' depends on the phosphorylation of acidic 'split proteins' [4].…”

Section: Discussionmentioning

confidence: 94%

“…Two of them were identified as L44 and L45 [18, 19, 211 and are phosphorylated in vitro by the casein kinase type I1 [21,221. Studies on the physiological role of the phosphorylation of ribosomal acidic proteins have been in progress for 10 years [23] and involved the yeast system. As shown in Ballesta's laboratory, the phosphorylation of L44, L44 and L45 might control the binding process of those proteins to ribosomal particle.…”

mentioning

confidence: 99%

Specfic protein kinase from Saccharomyces cerevisiae cells phosphorylating 60S ribosomal proteins

Pilecki

Grankowski

Jacobs

et al. 1992

European Journal of Biochemistry

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A protein kinase, specific for 60s ribosomal proteins, has been isolated from Saccharomyces cerevisiae cells, purified to almost homogeneity and characterized. The isolated enzyme is not related to other known protein kinases. Enzyme purification comprised three chromatography steps ; DEAEcellulose, phosphocellulose and heparin -Sepharose. SDSjPAGE analysis of the purified enzyme, indicated a molecular mass of around 71 kDa for the stained single protein band. The specific activity of the protein kinase was directed towards the 60s ribosomal proteins L44, L44, L45 and a 38 kDa protein. All the proteins are phosphorylated only at the serine residues. None of the 40s ribosomal proteins were phosphorylated in the presence of the kinase. For that reason we have named the enzyme the 60s kinase. An analysis of the phosphopeptide maps of acidic ribosomal proteins, phosphorylated at either the 60s kinase or casein kinase 11, showed almost identical patterns. Using the immunoblotting technique, the presence of the kinase has been detected in extracts obtained from intensively growing cells. These findings suggest an important role played by the 60s kinase in the regulation of ribosomal activity during protein synthesis. 20 years ago, when Kaltschmidt and Wittmann described the gel electrophoresis procedure for the separation and identification of Escherichia coli ribosomal proteins [l], the two acidic proteins L7 and L12 (12 kDa) of the 50s subunit have become one of the most studied ribosomal proteins with respect to their three-dimensional structure and biological function.The functional and physicochemical equivalents of the procaryotic L7 and L12 are two ribosomal proteins, L44 and L45, from the eucaryotic 60s subunit (for review see [2]). Contrary to in animals, the ribosomes of Saccharomyces cerevisiae cells contain a third low-molecular-mass acidic protein. That protein, named L44', can be separated on DEAEcellulose [3] and by an electrofocusing procedure in acrylamide gel [4]. All three proteins have molecular masses of about 13 kDa, belong to 'split proteins ' [5] The phosphorylation of the crude eucaryotic ribosomes was initially reported by Kabat [lo] and by Loeb et al. [ll]. Later studies by other authors have shown that mammalian ribosomes are associated with cyclic-AMP-independent protein kinases [12-151. One of the enzymes has turned out to be a casein kinase type I1 (CKII) which phosphorylates proteins L44 and L45 [14]. For the yeast ribosomes, it has been found that the majority of phosphoproteins, y-32P labelled in vivo [16 -191 correspond to the proteins phosphorylated in vitro with . Two of them were identified as L44 and L45 [18, 19, 211 and are phosphorylated in vitro by the casein kinase type I1 [21, 221. Studies on the physiological role of the phosphorylation of ribosomal acidic proteins have been in progress for 10 years [23] and involved the yeast system. As shown in Ballesta's laboratory, the phosphorylation of L44, L44 and L45 might control the binding process of those proteins to ribosom...

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“…They do not assemble onto preribosomes in the nucleolus but cycle between ribosomes and a cytosolic pool in numerous species including, Artemia salina, Saccharomyces cerevisiae (yeast), humans, and rats (7)(8)(9)(10)(11). Saenz-Robles et al (12) demonstrated quantitatively that exponentially growing yeast cells contain more 12-kDa P-proteins/ribosome than cells in the stationary phase of growth.…”

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confidence: 99%

Regulated Heterogeneity in 12-kDa P-protein Phosphorylation and Composition of Ribosomes in Maize (Zea mays L.)

Szick-Miranda

Bailey‐Serres

2001

Journal of Biological Chemistry

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Maize (Zea mays L.) possesses four distinct ϳ12-kDa P-proteins (P1, P2a, P2b, P3) that form the tip of a lateral stalk on the 60 S ribosomal subunit. RNA blot analyses suggested that the expression of these proteins was developmentally regulated. Western blot analysis of ribosomal proteins isolated from various organs, kernel tissues during seed development, and root tips deprived of oxygen (anoxia) revealed significant heterogeneity in the levels of these proteins. P1 and P3 were detected in ribosomes of all samples at similar levels relative to ribosomal protein S6, whereas P2a and P2b levels showed considerable developmental regulation. Both forms of P2 were present in ribosomes of some organs, whereas only one form was detected in other organs. Considerable tissue-specific variation was observed in levels of monomeric and multimeric forms of P2a. P2b was not detected in root tips, accumulated late in seed embryo and endosperm development, and was detected in soluble ribosomes but not in membrane-associated ribosomes that copurified with zein protein bodies of the kernel endosperm. The phosphorylation of the 12-kDa P-proteins was also developmentally and environmentally regulated. The potential role of P2 heterogeneity in P-protein composition in the regulation of translation is discussed.A complex of acidic ribosomal proteins (r-proteins) 1 forms a universally conserved lateral stalk on the large ribosomal subunit that facilitates the translocation phase of protein synthesis (1). In eukaryotes the structure is formed by a complex of acidic phosphoproteins. P0 (ϳ35 kDa), homologous to prokaryotic L10, interacts with 28 S rRNA to form the base of the stalk, and P1 and P2 (ϳ12 kDa), homologous to prokayotic L7/L12, are tethered as dimers to the stalk (2-5). P1 and P2 are structurally similar; each protein has three domains that include an ␣-helical N-terminal region and a central, flexible acidic hinge region followed by a highly conserved C terminus (E/KSD/ EDMGFG/SLD). The C-terminal region of P0 is structurally similar to 12-kDa P-proteins because it possesses the three domains of P1 and P2 (for review, see Ref. 6).The 12-kDa P-proteins are the only r-proteins found in multiple copies within the ribosome. They do not assemble onto preribosomes in the nucleolus but cycle between ribosomes and a cytosolic pool in numerous species including, Artemia salina, Saccharomyces cerevisiae (yeast), humans, and rats (7-11). demonstrated quantitatively that exponentially growing yeast cells contain more 12-kDa P-proteins/ribosome than cells in the stationary phase of growth. This suggests that the level of P-proteins in yeast ribosomes is affected by the metabolic state of the cell and possibly reflects the translational activity of the ribosome. The presence of these proteins in ribosomes has been shown to stimulate the eEF2-dependent GTPase activity of ribosomes (13-17), poly(U)-directed phenylalanine synthesis (14, 18), and eEF1A binding (19). Hence, modulation of the 12-kDa P-protein component of ribosomes may impar...

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the acidic ribosomal phosphoproteins from Saccharomyces cerevisiae

Cited by 46 publications

References 20 publications

Human acidic ribosomal phosphoproteins P0, P1, and P2: analysis of cDNA clones, in vitro synthesis, and assembly.

Human acidic ribosomal phosphoproteins P0, P1, and P2: analysis of cDNA clones, in vitro synthesis, and assembly.

Specfic protein kinase from Saccharomyces cerevisiae cells phosphorylating 60S ribosomal proteins

Regulated Heterogeneity in 12-kDa P-protein Phosphorylation and Composition of Ribosomes in Maize (Zea mays L.)

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