scholarly journals Topography of Escherichia coli ribosomal proteins. The order of reactivity of thiol groups*

1974 ◽  
Vol 143 (3) ◽  
pp. 599-606 ◽  
Author(s):  
Anastasia Bakardjieva ◽  
Robert R. Crichton
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Thiol Groups ◽  
Ribosomal Subunits

1. 30S and 50S ribosomal subunits of Escherichia coli were treated with N-[2,3-14C]-ethylmaleimide and iodo[14C]acetamide. 2. The proteins in the native subunits which reacted with the reagents were S1,‡ S2, S12, S13, S18, S21, L2, L5, L6, L10, L11, L15, L17, L20, L26+28 and L27. 3. Several proteins, such as S1, S12, S14, S18, L2, L6, L10, L11 and either L26 or 28, had thiol groups in an oxidized form and reacted to a greater extent after reduction with β-mercaptoethanol or dithiothreitol. 4. The total number of thiol groups in 30S and 50S subunits was determined as 16–17 and 26–27 respectively. The total number of thiol groups in each ribosomal protein was also determined. 5. The reaction of 30S and 50S subunits with iodoacetamide under several different conditions established the order of reactivity of thiol groups.

Ribosomal protein S14 is altered by two-step emetine resistance mutations in Chinese hamster cells

1983 ◽  
Vol 3 (2) ◽  
pp. 190-197
Author(s):  
J J Madjar ◽  
M Frahm ◽  
S McGill ◽  
D J Roufa
Keyword(s):  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Ribosomal Subunit ◽  
Chinese Hamster ◽  
Complementation Group ◽  
Resistance Mutations ◽  
Cho Cell ◽  
Ribosomal Subunits ◽  
40S Ribosomal Subunit ◽  
One Step

Four two-dimensional polyacrylamide gel electrophoresis systems were used to identify 78 Chinese hamster cell ribosomal proteins by the uniform nomenclature based on rat liver ribosomal proteins. The 40S ribosomal subunit protein affected by Chinese hamster ovary (CHO) cell one-step emetine resistance mutations is designated S14 in the standard nomenclature. To seek unambiguous genetic evidence for a cause and effect relationship between CHO cell emetine resistance and mutations in the S14 gene, we mutagenized a one-step CHO cell mutant and isolated second-step mutant clones resistant to 10-fold-higher concentrations of emetine. All of the highly resistant, two-step CHO cell mutants obtained displayed additional alterations in ribosomal protein S14. Hybridization complementation tests revealed that the two-step CHO cell emetine resistance mutants were members of the same complementation group defined by one-step CHO cell mutants, EmtB. Two-step mutants obtained from a Chinese hamster lung cell emetine-resistant clone belong to the EmtA complementation group. The two-step and EmtB mutants elaborated 40S ribosomal subunits, which dissociated to 32S and 40S core particles in buffers containing 0.5 M KCl at 4 degrees C. In contrast, 40S ribosomal subunits purified from all EmtA, one-step EmtB EmtC mutants, and wild-type CHO and lung cells were stable at this temperature in buffers containing substantially higher concentrations of salt. Thus, two-step emtB mutations affect the structure of S14 protein directly and the stability of the 40S ribosomal subunit indirectly.

scholarly journals The yeast omnipotent suppressor SUP46 encodes a ribosomal protein which is a functional and structural homolog of the Escherichia coli S4 ram protein.

Genetics ◽  
1992 ◽  
Vol 132 (2) ◽  
pp. 375-386 ◽  
Author(s):  
A Vincent ◽  
S W Liebman
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Protein ◽  
Dna Sequences ◽  
Ribosomal Proteins ◽  
Amino Acid Sequences ◽  
Ribosomal Protein Genes ◽  
Significant Homology ◽  
A Cell ◽  
Functional Equivalent ◽  
Ribosomal Protein S13

Abstract The accurate synthesis of proteins is crucial to the existence of a cell. In yeast, several genes that affect the fidelity of translation have been identified (e.g., omnipotent suppressors, antisuppressors and allosuppressors). We have found that the dominant omnipotent suppressor SUP46 encodes the yeast ribosomal protein S13. S13 is encoded by two similar genes, but only the sup46 copy of the gene is able to fully complement the recessive phenotypes of SUP46 mutations. Both copies of the S13 genes contain introns. Unlike the introns of other duplicated ribosomal protein genes which are highly diverged, the duplicated S13 genes have two nearly identical DNA sequences of 25 and 31 bp in length within their introns. The SUP46 protein has significant homology to the S4 ribosomal protein in prokaryotic-type ribosomes. S4 is encoded by one of the ram (ribosomal ambiguity) genes in Escherichia coli which are the functional equivalent of omnipotent suppressors in yeast. Thus, SUP46 and S4 demonstrate functional as well as sequence conservation between prokaryotic and eukaryotic ribosomal proteins. SUP46 and S4 are most similar in their central amino acid sequences. Interestingly, the alterations resulting from the SUP46 mutations and the segment of the S4 protein involved in binding to the 16S rRNA are within this most conserved region.

scholarly journals Ribosomal protein L30 is dispensable in the yeast Saccharomyces cerevisiae.

1990 ◽  
Vol 10 (10) ◽  
pp. 5235-5243 ◽  
Author(s):  
D M Baronas-Lowell ◽  
J R Warner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Wild Type ◽  
Diploid Strain ◽  
Chain Reaction ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ribosomal Subunits ◽  
Strong Homology ◽  
Polymerase Chain

In the yeast Saccharomyces cerevisiae, L30 is one of many ribosomal proteins that is encoded by two functional genes. We have cloned and sequenced RPL30B, which shows strong homology to RPL30A. Use of mRNA as a template for a polymerase chain reaction demonstrated that RPL30B contains an intron in its 5' untranslated region. This intron has an unusual 5' splice site, C/GUAUGU. The genomic copies of RPL30A and RPL30B were disrupted by homologous recombination. Growth rates, primer extension, and two-dimensional ribosomal protein analyses of these disruption mutants suggested that RPL30A is responsible for the majority of L30 production. Surprisingly, meiosis of a diploid strain carrying one disrupted RPL30A and one disrupted RPL30B yielded four viable spores. Ribosomes from haploid cells carrying both disrupted genes had no detectable L30, yet such cells grew with a doubling time only 30% longer than that of wild-type cells. Furthermore, depletion of L30 did not alter the ratio of 60S to 40S ribosomal subunits, suggesting that there is no serious effect on the assembly of 60S subunits. Polysome profiles, however, suggest that the absence of L30 leads to the formation of stalled translation initiation complexes.

Ribosomal protein L30 is dispensable in the yeast Saccharomyces cerevisiae

1990 ◽  
Vol 10 (10) ◽  
pp. 5235-5243
Author(s):  
D M Baronas-Lowell ◽  
J R Warner
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Wild Type ◽  
Diploid Strain ◽  
Chain Reaction ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ribosomal Subunits ◽  
Strong Homology ◽  
Polymerase Chain

In the yeast Saccharomyces cerevisiae, L30 is one of many ribosomal proteins that is encoded by two functional genes. We have cloned and sequenced RPL30B, which shows strong homology to RPL30A. Use of mRNA as a template for a polymerase chain reaction demonstrated that RPL30B contains an intron in its 5' untranslated region. This intron has an unusual 5' splice site, C/GUAUGU. The genomic copies of RPL30A and RPL30B were disrupted by homologous recombination. Growth rates, primer extension, and two-dimensional ribosomal protein analyses of these disruption mutants suggested that RPL30A is responsible for the majority of L30 production. Surprisingly, meiosis of a diploid strain carrying one disrupted RPL30A and one disrupted RPL30B yielded four viable spores. Ribosomes from haploid cells carrying both disrupted genes had no detectable L30, yet such cells grew with a doubling time only 30% longer than that of wild-type cells. Furthermore, depletion of L30 did not alter the ratio of 60S to 40S ribosomal subunits, suggesting that there is no serious effect on the assembly of 60S subunits. Polysome profiles, however, suggest that the absence of L30 leads to the formation of stalled translation initiation complexes.

The relationship between the spoT gene, the synthesis of stable RNA, ribosomal proteins, and the ββ′ subunits of RNA polymerase following a nutritional shiftup of Escherichia coli

1978 ◽  
Vol 56 (6) ◽  
pp. 528-533 ◽  
Author(s):  
Stephen M. Boyle ◽  
Frederick Chu ◽  
Nathan Brot ◽  
Bruce H. Sells
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
Rna Synthesis ◽  
Transient Increase ◽  
Spot Gene ◽  
The Relationship

The level of ppGpp and rates of synthesis of stable RNA, ribosomal protein, and the β and β′ subunits of RNA polymerase were measured following a nutritional shiftup in Escherichia coli strains, NF 929 (spoT+) and NF 930 (spoT'−). In the spoT+ strain, ppGpp levels decreased 50% within 2 min following shiftup, and the rates of synthesis of stable RNA, ribosomal proteins, and the β and β′ subunits of RNA polymerase increased with little or no lag. In contrast, in the spoT− strain, ppGpp levels transiently increased 40% during the first 6 min following shiftup. An inhibition in the rate of stable RNA synthesis and a delay in the increased synthesis of ribosomal proteins and β and β′ subunits occurred concurrently with the transient increase in ppGpp. In addition, the DNA-dependent synthesis in vitro of the β and β′ subunits of RNA polymerase was inhibited by physiological levels of ppGpp. Because of the timing and magnitude of the changes in ppGpp levels in the spoT− strain versus the timing when the new rates of stable RNA, ribosomal protein, and β and β′ subunits synthesis are reached, it is concluded that ppGpp is not the sole element regulating the expression of these genes.

scholarly journals The acidic ribosomal phosphoprotein of eukaryotes and its relationship to ribosomal proteins L7 and L12 of Escherichia coli

1978 ◽  
Vol 176 (2) ◽  
pp. 569-572 ◽  
Author(s):  
D P Leader ◽  
A A Coia
Keyword(s):  
Escherichia Coli ◽  
Ribosomal Protein ◽  
Ribosomal Proteins ◽  
E Coli ◽  
Data Support ◽  
Reported Data ◽  
Acidic Ribosomal Phosphoprotein

The acidic ribosomal phosphoprotein, Lgamma, of Krebs II ascites cells was further characterized and compared with proteins L7 and L12 of Escherichia coli. Ribosomal protein Lgamma was selectively removed from 60S sibosomal subunits by 50% ethanol and 1M-NH4Cl, and antibodies raised against protein Lgamma cross-reacted with E. coli protein L12 in immunodiffusion experiments. These and other, previously reported, data support the proposal that the uekaryotic counterpart of E. coli proteins L7 and L12 is phosphorylated.

Effect of RP51 gene dosage alterations on ribosome synthesis in Saccharomyces cerevisiae

1985 ◽  
Vol 5 (12) ◽  
pp. 3429-3435
Author(s):  
N Abovich ◽  
L Gritz ◽  
L Tung ◽  
M Rosbash
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Growth Rate ◽  
Ribosomal Protein ◽  
Dosage Compensation ◽  
Ribosomal Proteins ◽  
Gene Dosage ◽  
Translational Efficiency ◽  
Phenotypic Analysis ◽  
Ribosomal Subunits ◽  
Single Deletion

The Saccharomyces cerevisiae ribosomal protein rp51 is encoded by two interchangeable genes, RP51A and RP51B. We altered the RP51 gene dose by creating deletions of the RP51A or RP51B genes or both. Deletions of both genes led to spore inviability, indicating that rp51 is an essential ribosomal protein. From single deletion studies in haploid cells, we concluded that there was no intergenic dosage compensation at the level of mRNA abundance or mRNA utilization (translational efficiency), although phenotypic analysis had previously indicated a small compensation effect on growth rate. Similarly, deletions in diploid strains indicated that no strong mechanisms exist for intragenic dosage compensation; in all cases, a decreased dose of RP51 genes was characterized by a slow growth phenotype. A decreased dose of RP51 genes also led to insufficient amounts of 40S ribosomal subunits, as evidenced by a dramatic accumulation of excess 60S ribosomal subunits. We conclude that inhibition of 40S synthesis had little or no effect on the synthesis of the 60S subunit components. Addition of extra copies of rp51 genes led to extra rp51 protein synthesis. The additional rp51 protein was rapidly degraded. We propose that rp51 and perhaps many ribosomal proteins are normally oversynthesized, but the unassembled excess is degraded, and that the apparent compensation seen in haploids, i.e., the fact that the growth rate of mutant strains is less depressed than the actual reduction in mRNA, is a consequence of this excess which is spared from proteolysis under this circumstance.

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