Search PubMed⌕ Search

Biomedical subjects

A Favre

Publications and source records attributed to A Favre.

At least 127 records · Page 7Linked to original sources
PubMed1981

Photocrosslinking of thiolated aminoacyl-tRNA to ribosomal RNA and proteins.

tRNA has been converted to a form that can be photoactivated by chemical modification of some of the exposed cytidine residues to thio-4-uridine A certain percentage of the modified molecules can be charged and bound to the ribosome; thiolated fMet-tRNAfMet is bound to the P-site as shown by puromycin reactivity. Near the UV irradiation produces covalent crosslinks between total thiolated AA-tRNA or fMet-tRNAfMet and the ribosome. AA-tRNA becomes crosslinked to both 30S and 50S subunits but fMet-tRNAfMet to 50S subunits alone. In each case, crosslinking of tRNA was found to be not only to ribosomal proteins, but also to rRNA. The covalent complexes appear sufficiently stable to allow identification of the proteins or rRNA sequences involved.

Escherichia coli↗
PubMed1981

tRNA thiolated pyrimidines as targets for near-ultraviolet-induced synthesis of guanosine tetraphosphate in Escherichia coli.

Illumination with near-ultraviolet light triggers synthesis of ppGpp (guanosine 3'-diphosphate 5'-diphosphate) not only in growing Escherichia coli cells containing the putative chromophore 4-thiouridine in their tRNAs [Ramabhadran, T. V and Jagger, J. (1976) Proc. Natl Acad. Sci. USA, 73, 59--69], but also in nuv- cells which lack 4-thiouridine. The burst of ppGpp in nuv- cells is, however, induced exclusively by light of wavelengths shorter than 350 nm. Its maximum level is half that obtained in the parental strain. This ppGpp synthesis is also under the control of the relA gene, indicating that it is due to the accumulation of uncharged tRNAs. A candidate likely to trigger this effect is a 5-methylaminomethyl-2-thiouracil residues present in the first position of the anticodon loop of tRNAGlu, tRNALys and one tRNAGln isoacceptor. In conditions in vitro, this base is highly photoreactive at wavelengths shorter than 350 nm. Furthermore, near-ultraviolet-photomodified tRNAGlu and tRNALys become poor substrates of their acylation enzyme.

Escherichia coli↗
PubMed1980

Effect of freezing and thawing on the structure of turnip yellow mosaic virus.

The uncoating of turnip yellow mosaic virus in vitro induced by freezing and thawing has been investigated using a variety of biochemical techniques including the aminoacylation capacity of the viral RNA and the ability of the RNA to stimulate protein synthesis, as well as physico-chemical techniques such as sucrose gradient centrifugation and electron microscopy by negative staining. In particular a fluorescence test has been developed that can serve as a routine method to quantify the RNA liberated during the freeze-thaw process. Escape of the viral RNA is a highly cooperative phenomenon: it depends critically on the virus concentration during freezing and thawing. Increasing the ionic strength or including foreign proteins diminish the escape of the RNA. The RNA is not damaged by this treatment and its liberation occurs without disruption of the viral capsid.

Aminoacylation↗
PubMed1980

4-Thiouridine triggers both growth delay induced by near-ultraviolet light and photoprotection.

4-Thiouridine, a rare nucleoside present in Escherichia coli tRNAs, has been recently proposed to be the major chromophore leading to near-ultraviolet (315-400-nm)-induced growth delay. Here this is established by the isolation of mutants exhibiting a reduced growth delay. The selection procedure involves several successive cycles of 365-nm illumination of the cells in the stationary phase, followed by growth for two or three generations. After the eighth cycle, the level of 4-thiouridine in the culture decreases to 20% of the original level and all individual clones tested show a 4-thiouridine deficiency. One mutant exhibiting a complete lack of 4-thiouridine in its tRNAs has been characterized. In the dark the growth characteristics of the mutant and of the parental strain are indistinguishable. In contrast after near-ultraviolet illumination the nuv mutation abolishes the growth delay and considerably reduces the photoprotection efficiency.

Escherichia coli↗
PubMed1977

[4-Thiouridine and photoprotection in Escherichia coli K 12].

A high level of protection is observed in the Escherichia coli K 12 strain AB 1157 rec A 1 nuv+ whose transfer RNA contains 4-thiouridine. In contrast, the photoprotection level is low and observed at higher doses in a strain which differs from the former by a single mutation, nuv-, (lack of 4-thiouridine). This nucleoside is therefore an important chromophore leading to photoprotection. This conclusion is corroborated by the similarity of the action spectra for 8-13 link formation in tRNA and for photoprotection.

Cell Survival↗
PubMed1977

[Genetic localization of a mutation rendering the growth of E coli K12 insensitive to illumination at 365 nm].

The genotype of the Nop mutant recently isolated from the E. coli K 12 strain AB 1157 has been characterized. This mutant lacks 4-thiouridine in its tRNA and is much less susceptible to near ultraviolet-induced growth delay than wild type cells. This phenotype results from a single mutation called nuv which has been localized on the E. coli genetic map. nuv is found by conjugation to lie between the origins of injection of Hfr P4X and Hfr cavalli in the vicinity of the lac gene. Cotrans-duction with bacteriophage P1 more precisely maps nuv at 0.3 min. clockwise from tsx.

Conjugation, Genetic↗
PubMed1977

Role of divalent ions in folding of tRNA.

The native structure of tRNA is not achieved in low salt (4.5 mM Na+, 25 degrees C), but can be restored by addition of divalent ions. We have explored the structure of the central region in Escherichia coli tRNAfMet by absorption and emission spectroscopy of 4-thiouracil, and the structure of the anticodon loop in yeast tRNAPhe by fluorescence of the 'Y' base, versus the number of manganese ions bound to tRNA, which was derived from electron spin resonance. The fluorescence of the reduced 8-13 photoproduct (in which 4-thiouracil at position 8 is crosslinked to cytosine at position 13) was also analysed. In low salt (e.g. 4.5 mM Na+), the region of 4-thiouracil is affected strongly as the first eight Mn2+ bind to tRNA, whereas the fluorescence of the 'Y' base is affected only after four Mn2+ are bound. Considering the structural similarities of the two tRNAs, this suggests that the reorganisation brought about by divalent ions starts in the central region, the anticodon loop being affected later. The binding of divalent ions to each region starts together with its restructuration. Monovalent ions can substitute for divalent ions in this process, a 15 mM sodium concentration being equivalent to the binding of the first five Mn2+. If divalent ions are then added, even the first ones distribute themselves between both the central and the anticodon region. Alternatively, the renaturation may be achieved by monovalent ions only, implying that no sites exist whose occupancy by divalent ions is crucial for the native structure. These observations suggest that the role and means of divalent ion binding to tRNA are largely explainable in terms of a simple maganese-phosphate binding supplemented by electrostatic interaction with distant phosphates.

Anticodon↗
PubMed1977

[Mutants of Escherichia coli deficient in 4-thiouridine in which growth is insensitive to illumination at 365 nm].

Thirteen different mutants of E coli K12 selected for a reduced near ultra-violet induced growth delay have been isolated. The t-RNAs extracted from these clones have all a depressed content of 4-thiouridine. One of these mutants, called Nop has an almost negligible growth delay and completely lacks 4-thiouridine in its t-RNAs. Thus we provide genetic proofs that 4-thiouridine is the chromophore for growth delay.

Escherichia coli↗
PubMed1975

The secondary structure and poly(A) content of globin messenger RNA as a pure RNA and in polyribosome-derived ribonucleoprotein complexes.

The conformation in solution of duck and rabbit globin mRNA, and of the duck mRNA in the mRNA - protein particle, has been investigated by optical methods and also by the use of the dye ethidium bromide which becomes highly fluorescent when intercalated into the double-stranded regions of a nucleic acid. On the basis of the properties of this dye and on the ability of homopolyribonucleotides to form double-stranded structures we have, in addition, developed a simple and sensitive assay for the detection and quantitisation of sequences rich in a particular residue that may be present in an RNA chain. In solution, 45 to 60% of the nucleotides of duck globin nRNA were found to be in bihelical regions. A similar degree of secondary structure was found in rabbit globin mRNA (this paper), as well as in calf lens mRNA and mRNAs from ewe mammary gland (other results). All samples of globin mRNA examined in this work containeda sequence of poly(A), which has poly(U) binding properties similar to that of synthetic poly(a): no specific interaction between the poly(A) sequence and the rest of the molecules can be detected. The fraction of adenosine residues within these poly(A) segments represents 4% in rabbit mRNA and 8 to 9% in duck mRNA. An additional adenosine-rich segment interspersed with guanosine and possibly other residues, was also detected in one duck mRNA sample. The RNA in the duck mRNA - protein particle is also highly structured. The melting profile in the range of 20 to 65 degrees C is quite similar to that of free mRNA and the ability of ethidium bromide to intercalate is reduced to the extent of 70%. Yet the dichroic spectra of free and bound mRNA are significantly distinct. These data suggest that free and protein-bound mRNA May have a very similar degree of secondary structure but with distinct detailed conformation in bihelical regions (change in base tilting for example). Direct evidence has been obtained that proteins stick to the poly(A) segment in the particle since the fraction of adenosine residues detectable by our poly(u) titration procedure is reduced to 50% of that observed in the free mRNA.

Animals↗
PubMed1975

tRNA tertiary structure in solution as probed by the photochemically induced 8-13 cross-link.

The conformation of ten purified tRNAs from Escherichia Coli has been investigated by means of the photo-induced cross-linking of 4Srd8 and Cyd13, which is sensitive to the juxtaposition of the two bases. Three tRNAs photo-react abnormally slowly; tRNAPhe, tRNAMet/m a and tRNAVal/2; a comparison with normally reacting species suggests that base 47 (Urd or modified Urd) is involved in a tertiary interaction in Class I tRNAs with the triplet 8, 14, 28. The UGA suppressor tRNATrp photoreacts significantly slower than the wild type. Thus the single base change Gua 24 to Ade induces a conformational change that alters the rate constant for the cross-linking reaction.

Base Sequence↗