A set of compatible tac promoter expression vectors

Dykxhoorn, Derek M.; Pierre, Rebecca St.; Linn, Thomas

doi:10.1016/0378-1119(96)00289-2

Cited by 192 publications

(134 citation statements)

References 14 publications

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“…Plasmid pEXT22 (Dykxhoorn et al, 1996) and the norB : : lacZ fusion fragment from RUG7500 ( Fig. 1) were digested with XbaI and HindIII and ligated.…”

Section: Methodsmentioning

confidence: 99%

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cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

et al. 2008

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The ability of Neisseria gonorrhoeae to reduce nitric oxide (NO) may have important immunomodulatory effects on the host during infection. Therefore, a comprehensive understanding of the regulatory mechanism of the nitric oxide reductase gene (norB) needs to be elucidated. To accomplish this, we analysed the functional regions of the norB upstream region. The promoter contains an extended "10 motif (TGNTACAAT) that is required for high-level expression. Deletion and substitution analysis of the norB upstream region revealed that no sequence upstream of the "10 motif is involved in norB regulation under anaerobic conditions or in the presence of NO. However, replacement of a 29 bp inverted repeat sequence immediately downstream of the extended "10 motif gave high levels of aerobic expression of a norB : : lacZ fusion. Insertional inactivation of gonococcal nsrR, predicted to bind to this inverted repeat sequence, resulted in the loss of norB repression and eliminated NO induction capacity. Single-copy complementation of nsrR in trans restored regulation of both norB transcription and NorB activity by NO. In Escherichia coli, expression of a gonococcal nsrR gene repressed gonococcal norB; induction of norB occurred in the presence of exogenously added NO. NsrR also regulates aniA and dnrN, as well as its own expression. We also determined that Fur regulates norB by a novel indirect activation method, by preventing the binding of a gonococcal ArsR homologue, a second repressor whose putative binding site overlaps the Fur binding site.

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“…Plasmid pEXT22 (Dykxhoorn et al, 1996) and the norB : : lacZ fusion fragment from RUG7500 ( Fig. 1) were digested with XbaI and HindIII and ligated.…”

Section: Methodsmentioning

confidence: 99%

“…The norB : : lacZ fusion in RUG7500 was amplified and cloned into the low-copynumber plasmid pEXT22 (Dykxhoorn et al, 1996), to make pVI1. The entire nsrR gene, including 415 bp upstream of the ATG start codon, was cloned upstream of norB in pVI1 to make pVI18.…”

Section: Nsrr Regulation Of Norb In E Colimentioning

confidence: 99%

cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

et al. 2008

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“…coli strain JM109 containing the 54 -dependent transcriptional pspAp-lacZ fusion carried by the tetracycline-resistant plasmid pSJ1 (S. E. Jones and M.B., unpublished work), a derivative of pMR25 (gift of M. R. K. Alley), was used to measure PspF 1-275 activity in vivo. pPB1 derivatives containing the mutant forms of pspF was XbaI-HindIII-digested, and the mutant sequences were cloned into the chloramphenicol-resistant pACT3 plasmid (14) so as to express the wild-type or mutant PspF proteins under the control of a tac promoter.…”

Section: Methodsmentioning

confidence: 99%

The ATP hydrolyzing transcription activator phage shock protein F of Escherichia coli : Identifying a surface that binds σ ⁵⁴

Bordes

Wigneshweraraj

Schumacher

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

139

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Members of the protein family called ATPases associated with various cellular activities (AAA ؉ ) play a crucial role in transforming chemical energy into biological events. AAA ؉ proteins are complex molecular machines and typically form ring-shaped oligomeric complexes that are crucial for ATPase activity and mechanism of action. The Escherichia coli transcription activator phage shock protein F (PspF) is an AAA ؉ mechanochemical enzyme that functions to sense and relay the energy derived from nucleoside triphosphate hydrolysis to catalyze transcription by the 54 -RNA polymerase. Closed promoter complexes formed by the 54 -RNA polymerase are substrates for the action of PspF. By using a protein fragmentation approach, we identify here at least one 54 -binding surface in the PspF AAA ؉ domain. Results suggest that ATP hydrolysis by PspF is coupled to the exposure of at least one 54 -binding surface. This nucleotide hydrolysis-dependent presentation of a substrate binding surface can explain why complexes that form between 54 and PspF are transient and could be part of a mechanism used generally by other AAA ؉ proteins to regulate activity.

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“…To provide a compatible plasmid expressing additional controllable levels of DmpR, the KpnI site of the polylinker of the Sp R pEXT21 vector (IncW; three copies per cell; Dykxhoorn et al, 1996) was converted to an NdeI site via a linker to give pVI898. Subsequent cloning of the dmpR gene as an NdeI to HindIII fragment from pVI399 (Shingler and Moore, 1994) between these sites of pVI898 to give pVI899 places the dmpR gene cloned under the control of the lacI/Ptac promoter of the vector.…”

Section: Plasmid Constructionmentioning

confidence: 99%

The guanosine tetraphosphate (ppGpp) alarmone, DksA and promoter affinity for RNA polymerase in regulation of σ⁵⁴‐dependent transcription

Bernardo

Johansson

Solera

et al. 2006

Molecular Microbiology

130

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SummaryThe RNA polymerase-binding protein DksA is a cofactor required for guanosine tetraphosphate (ppGpp)-responsive control of transcription from s 70 promoters. Here we present evidence: (i) that both DksA and ppGpp are required for in vivo s 54 transcription even though they do not have any major direct effects on s 54 transcription in reconstituted in vitro transcription and s -factor competition assays, (ii) that previously defined mutations rendering the housekeeping s

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A set of compatible tac promoter expression vectors

Cited by 192 publications

References 14 publications

cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

The ATP hydrolyzing transcription activator phage shock protein F of Escherichia coli : Identifying a surface that binds σ ⁵⁴

The guanosine tetraphosphate (ppGpp) alarmone, DksA and promoter affinity for RNA polymerase in regulation of σ⁵⁴‐dependent transcription

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A set of compatible tac promoter expression vectors

Cited by 192 publications

References 14 publications

cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

cis- and trans-acting elements involved in regulation of norB (norZ), the gene encoding nitric oxide reductase in Neisseria gonorrhoeae

The ATP hydrolyzing transcription activator phage shock protein F of Escherichia coli : Identifying a surface that binds σ 54

The guanosine tetraphosphate (ppGpp) alarmone, DksA and promoter affinity for RNA polymerase in regulation of σ54‐dependent transcription

Contact Info

Product

Resources

About

The ATP hydrolyzing transcription activator phage shock protein F of Escherichia coli : Identifying a surface that binds σ ⁵⁴

The guanosine tetraphosphate (ppGpp) alarmone, DksA and promoter affinity for RNA polymerase in regulation of σ⁵⁴‐dependent transcription