The yeast 5S rRNA binding robosomal protein YL3 is phosphorylated in vivo

Campos, Francisco; Corona-Reyes, M.; Zínker, Samuel

doi:10.1016/0167-4781(90)90198-b

Cited by 10 publications

(7 citation statements)

References 25 publications

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“…None of the rpL1-HA point mutations were predicted to disrupt its major structural elements (data not shown). Previous studies have indicated that rpL1 is phosphorylated in vivo; up to two residues are thought to be phosphorylated per mol of rpL1 (Zinker and Warner, 1976;Campos et al, 1990). Although the significance of this phosphorylation is not yet known, it is possible that the rpL1-HA-2 (T28A) mutant protein and the rpL1-HA-6 and rpL1-HA-7 proteins that contain the carboxylterminal deletion are defective because they alter potential phosphorylation sites and thus affect rpL1 function.…”

Section: Discussionmentioning

confidence: 99%

Multiple Regions of Yeast Ribosomal Protein L1 Are Important for Its Interaction with 5 S rRNA and Assembly into Ribosomes

Deshmukh

Stark

Yeh³

et al. 1995

Journal of Biological Chemistry

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Yeast ribosomal protein L1 binds to 5 S rRNA and can be released from 60 S ribosomal subunits as an intact ribonucleoprotein particle. To identify residues important for binding of Saccharomyces cerevisiae rpL1 to 5 S rRNA and assembly into functional ribosomes, we have isolated mutant alleles of the yeast RPL1 gene by site-directed and random mutagenesis. The rpl1 mutants were assayed for association of rpL1 with 5 S rRNA in vivo and in vitro and assembly of rpL1 into functional 60 S ribosomal subunits. Consistent with previous data implicating the importance of the carboxyl-terminal 47 amino acids of rpL1 for binding to 5 S rRNA in vitro, we find that deletion of the carboxyl-terminal 8, 25, or 44 amino acids of rpL1 confers lethality in vivo. Missense mutations elsewhere in rpL1 also affect its function, indicating that multiple regions of rpL1 are important for its association with 5 S rRNA and assembly into ribosomes.

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Section: Discussionmentioning

confidence: 99%

Multiple Regions of Yeast Ribosomal Protein L1 Are Important for Its Interaction with 5 S rRNA and Assembly into Ribosomes

Deshmukh

Stark

Yeh³

et al. 1995

Journal of Biological Chemistry

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“…Afterwards, protein P0 was also characterized as a phoshoprotein immunologically related to proteins P1 and P2 [7]. More recently, the 5S rRNA binding protein L3 was reported to be phosphorylated in yeast [8].…”

Section: Introductionmentioning

confidence: 99%

Phosphorylation of the yeast ribosomal stalk. Functional effects and enzymes involved in the process

et al. 1999

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The ribosomal stalk is directly involved in the interaction of the elongation factors with the ribosome during protein synthesis. The stalk is formed by a complex of five proteins, four small acidic polypeptides and a larger protein which directly interacts with the rRNA at the GTPase center. In eukaryotes the acidic components correspond to the 12-kDa P1 and P2 proteins, and the RNA binding component is the P0 protein. All these proteins are found phosphorylated in eukaryotic organisms, and previous in vitro data suggested this modification was involved in the activity of this structure. Results from mutational studies have shown that phosphorylation takes place at a serine residue close to the carboxy end of the P proteins. Modification of this serine residue does not affect the formation of the stalk and the activity of the ribosome in standard conditions but induces an osmoregulation-related phenotype at 37 degrees C. The phosphorylatable serine is part of a consensus casein kinase II phosphorylation site. However, although CKII seems to be responsible for part of the stalk phosphorylation in vivo, it is probably not the only enzyme in the cell able to perform this modification. Five protein kinases, RAPI, RAPII and RAPIII, in addition to the previously reported CKII and PK60 kinases, are able to phosphorylate the stalk proteins. A comparison of the five enzymes shows differences among them that suggest some specificity regarding the phosphorylation of the four yeast acidic proteins. It has been found that some typical effectors of the PKC kinase stimulate the in vitro phosphorylation of the stalk proteins. All the data suggest that although phosphorylation is not involved in the interaction of the acidic P proteins with the ribosome, it can affect the ribosome activity and might participate in a possible ribosome regulatory mechanism.

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“…We did not observe any modifications caused by phosphorylation. It has been well established that the acidic ribosomal proteins (rpP0, rpP1A, rpP1B, rpP2A, and rpP2B) are phosphorylated in yeast (24,48), but only a single report has identified up to two serine phosphorylation sites in a core large subunit protein (rpL5) (49). As mentioned earlier, the yeast acidic ribosome subcomplex dynamically associates with the large subunit (16,24,42), and the apparent absence of the acidic ribosomal proteins is likely caused by their absence in the original sample, rather than difficulties encountered during the reversed-phase separation or ESI.…”

Section: N-terminal Modificationsmentioning

confidence: 99%

Direct mass spectrometric analysis of intact proteins of the yeast large ribosomal subunit using capillary LC/FTICR

Lee

Berger

Martinović

et al. 2002

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

183

133

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Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry coupled with capillary reverse-phase liquid chromatography was used to characterize intact proteins from the large subunit of the yeast ribosome. High mass measurement accuracy, achieved by ''mass locking'' with an internal standard from a dual electrospray ionization source, allowed identification of ribosomal proteins. Analyses of the intact proteins revealed information on cotranslational and posttranslational modifications of the ribosomal proteins that included loss of the initiating methionine, acetylation, methylation, and proteolytic maturation. High-resolution separations permitted differentiation of protein isoforms having high structural similarity as well as proteins from their modified forms, facilitating unequivocal assignments. The study identified 42 of the 43 core large ribosomal subunit proteins and 58 (of 64 possible) core large subunit protein isoforms having unique masses in a single analysis. These results demonstrate the basis for the high-throughput analyses of complex mixtures of intact proteins, which we believe will be an important complement to other approaches for defining protein modifications and their changes resulting from physiological processes or environmental perturbations.M ass spectrometry has evolved into a powerful tool for analyzing biomolecules because of the development of matrix-assisted laser desorption ionization (1, 2) and electrospray ionization (ESI) (3) and advances in both mass analyzers and data processing capabilities. With these ionization methods, which are amenable to megadalton-size molecules, the broadly useful detection and characterization of biopolymers by mass spectrometric methods are feasible. For example, MS has become a preferred analytical tool for proteome analyses, in which dynamic populations of proteins are identified and quantified. Proteins are now routinely identified by mass spectrometric and tandem mass spectrometric analysis of proteolytic digests of individual protein spots on two-dimensional (2D) PAGE (4). However, the intact protein level analyses of complex protein mixtures, while potentially providing complementary and direct information for protein identification, have been a much greater experimental challenge and only rarely attempted (5, 6).Many pivotal cellular processes are not carried out by individual proteins, but rather by large protein-protein complexes and protein-nucleic acid complexes. The analyses of such complexes have been greatly expanded by improvements in both biological separations and MS. One of the more complicated protein complexes yet studied by MS is the cytosolic ribosome complex. Efforts to define the protein composition (7, 8), modifications (9), and subunit interactions (10) of ribosomes generally have mirrored the development of biomolecular analysis techniques and experimental approaches for the study of noncovalent protein complexes.Ribosomes are the canonical example of ribonucleoprotein complexes in both prokaryot...

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The yeast 5S rRNA binding robosomal protein YL3 is phosphorylated in vivo

Cited by 10 publications

References 25 publications

Multiple Regions of Yeast Ribosomal Protein L1 Are Important for Its Interaction with 5 S rRNA and Assembly into Ribosomes

Multiple Regions of Yeast Ribosomal Protein L1 Are Important for Its Interaction with 5 S rRNA and Assembly into Ribosomes

Phosphorylation of the yeast ribosomal stalk. Functional effects and enzymes involved in the process

Direct mass spectrometric analysis of intact proteins of the yeast large ribosomal subunit using capillary LC/FTICR

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