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L A Isaksson

Publications and source records attributed to L A Isaksson.

At least 55 records · Page 3Linked to original sources
PubMed1986

Influence of modification next to the anticodon in tRNA on codon context sensitivity of translational suppression and accuracy.

Effects on translation in vivo by modification deficiencies for 2-methylthio-N6-isopentenyladenosine (ms2i6A) (Escherichia coli) or 2-methylthio-N6-(4-hydroxyisopentenyl)adenosine (ms2io6A) (Salmonella typhimurium) in tRNA were studied in mutant strains. These hypermodified nucleosides are present on the 3' side of the anticodon (position 37) in tRNA reading codons starting with uridine. In E. coli, translational error caused by tRNA was strongly reduced in the case of third-position misreading of a tryptophan codon (UGG) in a particular codon context but was not affected in the case of first-position misreading of an arginine codon (CGU) in another codon context. Misreading of UGA nonsense codons at two different positions was codon context dependent. The efficiencies of some tRNA nonsense suppressors were decreased in a tRNA-dependent manner. Suppressor tRNA which lacks ms2i6A-ms2io6A becomes more sensitive to codon context. Our results therefore indicate that, besides improving translational efficiency, ms2i6A37 and ms2io6A37 modifications in tRNA are also involved in decreasing the intrinsic codon reading context sensitivity of tRNA. Possible consequences for regulation of gene expression are discussed.

Anticodon↗
PubMed1985

Involvement of ribosomal protein L7/L12 in control of translational accuracy.

The effects of two mutations, which map at the rplL locus and both give a changed 50S ribosomal protein L7/L12, were studied. Both mutations are associated with an increased misreading of all three nonsense codons in vivo and ribosomes from the mutants give an increased misreading of the phenylalanine codon UUU by tRNALeu in vitro. The rplL-associated misreading in vitro is not limited to a particular type of mRNA or tRNA. Results from a translational proofreading assay, using mutant ribosomes, suggest that protein L7/L12 is involved in the control of translational accuracy by contributing to the efficiency of a translational proofreading step(s).

Bacterial Proteins↗
PubMed1984

A temperature-sensitive mutant of Escherichia coli that shows enhanced misreading of UAG/A and increased efficiency for some tRNA nonsense suppressors.

A spontaneous mutant was isolated that harbors a weak suppressing activity towards a UAG mutation, together with an inability to grow at 43 degrees C in rich medium. The mutation is shown to be associated with an increased misreading of UAG at certain codon contexts and UAA. UGA, missense or frameshift mutations do not appear to be misread to a similar extent. The mutation gives an increased efficiency to several amber tRNA suppressors without increasing their ambiguity towards UAA. The ochre suppressors SuB and Su5 are stimulated in their reading of both UAG and UAA with preference for UAG. An opal suppressor is not affected. The effect of the mutation on the efficiency of amber and ochre suppressors is dependent on the codon context of the nonsense codon. The mutated gene (uar) has been mapped and found to be recessive both with respect to suppressor-enhancing ability as well as for temperature sensitivity. The phenotype is partly suppressed by the ochre suppressor SuC. It is suggested that uar codes for a protein, which is involved in translational termination at UAG and UAA stop codons.

Chromosomes, Bacterial↗
PubMed1982

Translation rates and misreading characteristics of rpsD mutants in Escherichia coli.

Three ribosomal ambiguity (Ram) mutants, changed in ribosomal protein S4, have been examined with respect to elongation rate and misreading of translation in vivo and in vitro. Ram mutants increase misreading of nonsense codons in vivo, compared to wild type, between 2-50 times depending on the nature of the nonsense codon, its position, and which rpsD allele is present. Ram ribosomes also show an increased error frequency in vitro. The elongation rate of translation does not seem to be significantly changed, neither in vivo nor in vitro, irrespective of which rpsD allele is present. We suggest that there exists no general relationship between the accuracy and the overall speed of translation in Ram strains.

Alleles↗
PubMed1980

Effects of growth conditions and mutations in RNA polymerase on translational activity in vitro in Escherichia coli.

The translational capacity in vitro in Escherichia coli, using RNA from phage R17 or Q beta as messenger, is several times higher if the extracts are prepared from cells harvested in early exponential phase or grown under conditions of good aeration compared to if extracts are prepared from cells harvested in a later growth phase or grown under semi-aerobic conditions. In low activity extracts the production of phage replicase protein is preferentially affected. Growth of a wild type strain under semi-aerobic conditions has a less pronounced effect on translational capacity in vitro using crude mRNA from normal or T4 infected cells or with poly(U). Mutants were fortuitously found which did not show a lowered translational activity in vitro as a result of entering late phase of growth. Two of these were changed in RNA polymerase. Two different translational inhibitors can be demonstrated in the ribosomal wash fraction obtained from semi-aerobically grown wild type cells, whereas only one was found in the case of aerobically grown cells. The low translational activity of semi-aerobically grown cells in vitro is implied to be dependent on the induction or activation of a translational inhibitor. It behaves like a protein but is not likely to be a protease or RNAse.

Bacteriophages↗
PubMed1980

Analysis of rpsD mutations in Escherichia coli. IV. Accumulation of minor forms of protein S7(K) in ribosomes of rpsD mutant strains due to translational read-through.

A certain proportion of protein S7 exists in an altered form in E. coli rpsD (S4) mutants. Depending on the type of S4 mutation involved, two different forms of the altered S7 can be distinguished. The unusual form is longer than normal S7 by about 500 daltons due to extra material at the carboxyl end of the protein. It is suggested that a mutationally altered S4 might lower the efficiency of termination during translation of the messenger for S7. This results in an increased frequency of translational read-through, which gives the observed longer forms of S7. Data are interpreted to mean that one class of S4 mutants might suppress UGA and UAG whereas another class only suppresses UGA.

Amino Acids↗
PubMed1979

Analysis of rpsD mutations in Escherichia coli. I. Comparison of mutants with various alterations in ribosomal protein S4.

Streptomycin-independent revertants were selected from streptomycin-dependent mutants. Twenty-five out of 150 such revertants were temperature sensitive. Ribosomal proteins from 18 temperature-sensitive and 10 temperature-insensitive revertants were analysed by SDS-polyacrylamide gel electrophoresis. Seventeen of the former but none of the latter category showed an alteration of protein S4. The mutated rpsD allele of 6 temperature-sensitive revertants was transduced into a rpsL+ strain. In all cases an increased suppressibility of T4 amber phages was observed. Such suppressibility was not observed in the original rpsD, rpsL strains. All 18 temperature-sensitive mutants were disturbed in the processing of 17s to 16s RNA at non-permissive temperature and the accumulated 17s RNA was degraded. Temperature-insensitive rpsD revertants could be isolated, which had gained a second alteration in S4. Such revertants, which had lost the temperature-sensitive property, were also unable to suppress growth of T4 amber phages. It is concluded that temperature-sensitive growth, inability to process 17s RNA and to assemble 30S ribosomes at non-permissive temperature as well as increased translational ambiguity are highly correlated properties in rpsD mutants.

Bacterial Proteins↗
PubMed1979

Analysis of rpsD mutations in Escherichia coli. III. Effects of rpsD mutations on expression of some ribosomal protein genes.

Relative rates of production and steady state levels of ribosomal proteins were determined in a temperature sensitive rpsD (S4) mutant of Escherichia coli. Some proteins (S4, S12, S13) were overproduced in the mutant at permissive temperature but steady state levels of all examined ribosomal proteins were normal. In a rpsD+/rpsD+ homodiploid strain the relative rates of production of ribosomal proteins were not affected by the increased gene dose. In a rpsD+/rpsD heterodiploid strain only wild type, but not mutant S4, was found. In such a strain S4, S7, S12 and probably S13 is overproduced. It is implied that S4 is involved in the regulation of expression of proximal genes of the two transcriptional units including the genes coding for S4 itself and S12, respectively. A degradation system for ribosomal proteins, which is rapid enough to be of regulatory significance, is demonstrated.

Bacterial Proteins↗
PubMed1978

Mutations in the tryptophanyl-transfer ribonucleic acid ligase of E. coli causing temperature-sensitivity for growth.

Strains of Escherichia coli K-12 with altered tryptophanyl-tRNA ligases, conferring temperature-sensitivity for growth, have been isolated as spontaneous one-step mutants. The mutated enzymes differ markedly in activity from the wildtype, even at the permissive temperature. When assayed at the non-permissive temperature, the mutant enzymes are completely inactive. In one of the mutant strains, growth can be completely inhibited by addition of L-phenylalanine or L-tyrosine to the medium.

Amino Acyl-tRNA Synthetases↗
PubMed1978

The temperature sensitive mutant 72c. I. Pleiotropic growth behaviour and changed response to some antibiotics and mutations in the transcription or translation apparatus.

The spontaneous temperature sensitive mutant 72c is shown to be more tolerant to fusidic acid, but less tolerant to trimethoprim on plates at permissive temperature, than is the parental strain. The poor growth of the mutant on amino acids supplemented plates, as well as its inability to grow on broth plates at 40 degrees, can be compensates by sublethal amounts of chloroamphenicol. Also some mutations to Rif-R or Str-R improve growth of the mutant under certain conditions. Reversion and other genetic analysis strongly suggest, that the pleiotropic behaviour of the mutant is due to a single mutation in a gene, which is designated fusB and is closely cotransducible with lip at min 14 of the E. coli chromosome. The gene order is lip-fusB-supE.

Chloramphenicol↗
PubMed1978

The temperature sensitive mutant 72c. II. Accumulation at high temperature of ppGpp and pppGpp in the presence of protein synthesis.

A heat sensitive mutant of E. coli has been analyzed. A shift to restrictive temperature leads to an accumulation of ppGpp and pppGpp in both the parental and the mutant strains (both are relA+). The pool of these compounds is shown to decrease with time after the temperature shift in the case of the parental strain, but remains at the same elevated level in the case of the mutant. The temperature shift of the mutant leads to an apparent reduction of stable RNA synthesis; this inhibition can be released by chloroamphenicol or tetracycline. Gross protein synthesis is more or less unaffected at restrictive temperature. In the parental strain little effect is seen on RNA and protein synthesis after the temperature shift. A relA derivative of the mutant does not show the same inhibition of RNA synthesis at high temperature. Sedimentation analysis suggests that mutant 70S ribosome are more stable, when exposed to a lowered Mg2+ concentration, than are 70S ribosomes from the parental strain. In addition, the relative amounts of the two forms of ribosomal protein S6, which can be obtained on DEAE chromatography (Held et al., 1973), are significantly changed in the mutant.

Bacterial Proteins↗
PubMed1978

Temperature-sensitive Escherichia coli mutant producing a temperature-sensitive sigma subunit of DNA-dependent RNA polymerase.

A gene affecting the sigma subunit of DNA-dependent RNA polymerase is tightly linked to dnaG at 66 min on the Escherichia coli chromosome. In order to create an easily selectable marker in this region, we inserted transposon-10, which carries a gene determining resistance to tetracycline (tet) near 66 min, and the order tolC-dnaG-sigma-tet was determined. We used frequency of contransduction with tet as a criterion to screen a collection of spontaneous temperature-sensitive Escherichia coli mutants that might affect the sigma subunit. One such mutant was found to map at the sigma locus. The sigma subunit isolated from this mutant is unstable at 46 degrees C in vitro and has an altered electrophoretic mobility. The temperature sensitivity of RNA synthesis in this mutant indicates that most transcription in E. coli is sigma dependent.

Chromosome Mapping↗
PubMed1977

A procedure for isolation of spontaneous mutants with temperature sensitive of RNA and/or protein.

A procedure for the isolation of spontaneous temperature sensitive mutants of Escherichia coli has been developed. They are selected as survivors at high temperature against the combined killing effects exerted by a temperature inducible lambda prophage and either streptomycin plus ampillicin or ampicillin plus cycloserine. The mutants so obtained are blocked in vivo in the synthesis of RNA or protein or both at restrictive temperature.

Bacterial Proteins↗