| 1. |
- Hughes, Diarmaid, 1956-
(author)
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Both genes for EF-Tu in Salmonella typhimurium are individually dispensible for growth
- 1990
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In: Journal of Molecular Biology. - 0022-2836 .- 1089-8638. ; 215:1, s. 41-51
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Journal article (peer-reviewed)abstract
- Each of the two genes encoding EF-Tu in Salmonella typhimurium has been inactivated using a mini-Mu MudJ insertion. Eleven independently isolated insertions are described, six in tufA and five in tufB. Transduction analysis shows that the inserted MudJ is 100% linked to the appropriate tuf gene. A mutant strain with electrophoretically distinguishable EF-TuA and EF-TuB was used to show, on two-dimensional gels, that the MudJ insertions result in the loss of the appropriate EF-Tu protein. Southern blotting, using cloned Escherichia coli tuf sequences as probes, shows that each MudJ insertion results in the physical breakage of the appropriate tuf gene. The degree of growth-rate impairment associated with each tuf inactivation is independent of which tuf gene is inactivated. The viability of S. typhimurium strains with either tuf gene inactive contrasts strongly with data suggesting that in the closely related bacterium E. coli, an active tufA gene is essential for growth. Finally the strains described here facilitate the analysis of phenotypes associated with individual mutant or wild-type Tus both in vivo and in vitro.
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| 2. |
- Abdulkarim, Farhad, et al.
(author)
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Homologous recombination between the tuf genes of Salmonella typhimurium
- 1996
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In: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 260:4, s. 506-522
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Journal article (peer-reviewed)abstract
- The genes coding for the translation factor EF-Tu, tufA and tufB are separated by over 700 kb on the circular chromosome of Salmonella typhimurium. The coding regions of these genes have 99% identity at the nucleotide level in spite of the presumed ancient origin of the gene duplication. Sequence comparisons between S. typhimurium and Escherichiacoli suggest that within each species the two tuf genes are evolving inconcert. Here we show that each of the S. typhimurium tuf genes cantransfer genetic information to the other. In our genetic system thetransfers are seen as non-reciprocal, i.e. as gene conversion events.However, the mechanism of recombination could be reciprocal, with sisterchromosome segregation and selection leading to the isolation of aparticular class of recombinant. The amount of sequence informationtransferred in individual recombination events varies, but can be close tothe entire length of the gene. The recombination is RecABCD-dependent,and is opposed by MutSHLU mismatch repair. In the wild-type, this typeof recombination occurs at a rate that is two or three orders of magnitudegreater than the nucleotide substitution rate. The rate of recombinationdiffers by six orders of magnitude between a recA and a mutS strain.Mismatch repair reduces the rate of this recombination 1000-fold. The rateof recombination also differs by one order of magnitude depending onwhich tuf gene is donating the sequence selected for. We discuss threeclasses of model that could, in principle, account for the sequencetransfers: (1) tuf mRNA mediated recombination; (2) non-allelic reciprocalrecombination involving sister chromosomes; (3) non-allelic geneconversion involving sister chromosomes, initiated by a double-strandbreak close to one tuf gene. Although the mechanism remains to bedetermined, the effect on the bacterial cells is tuf gene sequencehomogenisation. This recombination phenomenon can account for theconcerted evolution of the tuf genes.
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| 3. |
- Arwidsson, Ola, et al.
(author)
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Evidence against reciprocal recombination as the basis for tuf gene conversion in Salmonella enterica serovar Typhimurium
- 2004
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In: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 338:3, s. 463-467
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Journal article (peer-reviewed)abstract
- The duplicate tuf genes on the Salmonella enterica serovar Typhimurium chromosome co-evolve by a RecA-, RecB-dependent gene conversion mechanism. Gene conversion is defined as a non-reciprocal transfer of genetic information. However, in a replicating bacterial chromosome there is a possibility that a reciprocal genetic exchange between different tuf genes sitting on sister chromosomes could result in "apparent" gene conversion. We asked whether the major mechanism of tuf gene conversion was classical or apparent. We devised a genetic selection that allowed us to isolate and examine both expected products from a reciprocal recombination event between the tuf genes. Using this selection we tested within individual cultures for a correlation in the frequency of jackpots as expected if recombination were reciprocal. We found no correlation, either in the frequency of each type of recombinant product, or in the DNA sequences of the products resulting from each recombination event. We conclude that the evidence argues in favor of a non-reciprocal gene conversion mechanism as the basis for tuf gene co-evolution.
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| 4. |
- Hughes, Diarmaid, 1956-
(author)
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Co-evolution of the tuf genes links gene conversion with the generation of chromosomal inversions
- 2000
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In: Journal of Molecular Biology. - : ACADEMIC PRESS LTD. - 0022-2836 .- 1089-8638. ; 297:2, s. 355-364
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Journal article (peer-reviewed)abstract
- The tufA and tufB genes in Salmonella typhimurium co-evolve by recombination and exchange of genetic material. A model is presented which predicts that co-evolution is achieved by gene conversions and chromosomal inversions. Analysis of recombinants reveals that conversion and inversion each occur with similar rates and each depends on RecBCD activity. The model predicts sequence structures for different classes of post-recombination tuf genes. Sequence analysis reveals the presence of each of these structures and classes, with a predicted bias in the absence of mismatch repair. An implication of these data is that co-evolution of gene families can be linked with the generation of chromosomal rearrangements.
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| 5. |
- Laurberg, M, et al.
(author)
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Structure of a mutant EF-G reveals domain III and possibly the fusidic acid binding site
- 2000
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In: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 303:4, s. 593-603
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Journal article (peer-reviewed)abstract
- The crystal structure of Thermus thermophilus elongation factor G (EF-G) carrying the point mutation His573Ala was determined at a resolution of 2.8 Å. The mutant has a more closed structure than that previouslyreported for wild-type EF-G. This is obtained by a 10° rigid rotation of domains III, IV and V with regardto domains I and II. This rotation results in a displacement of the tipof domain IV by approximately 9 Å. The structure of domain III is nowfully visible and reveals the double split β-α-β motif also observed for EF-G domain V and for several ribosomal proteins. A large number of fusidic acid resistant mutations found in domain III have now been possible tolocate. Possible locations for the effector loop and a possible bindingsite for fusidic acid are discussed in relation to some of the fusidic acid resistant mutations.
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