| 1. |
- Smith, C A, et al.
(författare)
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The trfA and trfB promoter regions of broad host range plasmid RK2 share common potential regulatory sequences.
- 1984
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Ingår i: Nucleic Acids Research. - 0305-1048 .- 1362-4962. ; 12:8
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Tidskriftsartikel (refereegranskat)abstract
- The positions of the trfA and trfB promoters of broad host range IncP plasmid RK2 (identical to RP1, RP4, R68 and R18 ) were identified by RNA polymerase protection studies, and the nucleotide sequences of the promoter regions determined. A mutation within the trfA promoter sequence is associated with loss of kilD activity. In addition a probable promoter region for the kilB locus was identified. The three promoter regions share common palindromic sequences which may serve as sites for the coordinate regulation of replication and kil functions.
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| 2. |
- Shinger, V, et al.
(författare)
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Transcription in the trfA region of broad host range plasmid RK2 is regulated by trfB and korB.
- 1984
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Ingår i: Molecular General Genetics. - 0026-8925 .- 1432-1874. ; 195:3
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Tidskriftsartikel (refereegranskat)abstract
- Transcription at various points in the trf A region of broad host range plasmid RK2 has been analysed by measuring expression of the galK gene inserted at EcoRI sites introduced previously by TB1723 transposition mutagenesis. Rightward transcription (anti-clockwise on RK2) probably from a single promoter, proceeds across two open reading frames coding for a 13 kD polypeptide of unknown function, and the trf A gene, which provides a protein(s) essential for plasmid replication. This transcription is not auto-regulated by the products of either open reading frame and is also not subject to significant attenuation prior to the end of the trfA open reading frame. Leftward transcription appears to be directed by at least two well separated promoters, the more leftward being three to four times stronger than the more rightward. Rightward, but not leftward, transcription is repressed about 9-fold by the trfB locus of RK2 alone (so far not separable from the loci korA and korD) in trans while the combination of the korB and trfB loci in trans represses both rightward transcription (about 100-fold) and leftward transcription (the stronger activity by 10 to 15-fold). Regulation of these operons is therefore qualitatively different. The kilD locus in the trfA region, which is suppressed by korD (trfB) is thus probably part of the rightward (trfA) operon, while leftward transcription may represent the start of an operon containing kilB. The results suggest that RK2kor loci act by repressing transcription of kil loci and that the kil and kor control circuits may be part of an interlocking system of RK2 genes involved in replication and stable maintenance.
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| 3. |
- Shingler, V, et al.
(författare)
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Analysis of the trfA region of broad host-range plasmid RK2 by transposon mutagenesis and identification of polypeptide products.
- 1984
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Ingår i: Journal of Molecular Biology. - 0022-2836 .- 1089-8638. ; 175:3
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Tidskriftsartikel (refereegranskat)abstract
- Broad host-range plasmid RK2 is a member of the Escherichia coli incompatibility group P. Unlike most other groups of plasmids, members of the P group are capable of efficient transfer between and maintenance in most gram-negative bacterial species. It is of interest whether this broad host-range results from differences between the mechanism of replication of broad and narrow host-range plasmids. The regions of RK2 required for replication in E. coli have previously been defined as an origin of vegetative replication, oriVRK2 , and a gene, trfA , specifying a positively required trans-acting product. In this study Tn1723 transposon insertions have been used to map the trfA gene and determine its functional gene product. The Tn1723 insertions define the outer limits of the gene, a promoter region, a "leader" region not essential for trfA activity and a coding region. Three polypeptides of 13 X 10(3), 43 X 10(3) and 32 X 10(3) molecular weight are produced from this region and the production of a 32 X 10(3) Mr polypeptide is shown to be correlated with trfA activity in E. coli. Analysis of polypeptides produced from transposon insertion derivatives in which all but 35 base-pairs of inserted DNA is deleted, along with the effect of these insertions on trfA activity, suggest that the 43 X 10(3) and 32 X 10(3) Mr polypeptide coding sequences overlap in the same reading frame and that all three polypeptides (13 X 10(3), 32 X 10(3) and 43 X 10(3) Mr) may be translated from the same initial transcript.
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