Repressor Forms of the Enhancer-binding Protein NtrC: Some Fail in Coupling ATP Hydrolysis to Open Complex Formation by σ54-Holoenzyme

1996 ◽  
Vol 260 (3) ◽  
pp. 317-331 ◽  
Author(s):  
Anne K. North ◽  
David S. Weiss ◽  
Hideyuki Suzuki ◽  
Yehuda Flashner ◽  
Sydney Kustu
Keyword(s):  
Complex Formation ◽  
Atp Hydrolysis ◽  
Binding Protein ◽  
Enhancer Binding Protein ◽  
Open Complex Formation

scholarly journals Mechanism of Antiactivation at the Pseudomonas sp. Strain ADP σN-Dependent PatzTPromoter

2016 ◽  
Vol 82 (14) ◽  
pp. 4350-4362 ◽  
Author(s):  
Ana Isabel Platero ◽  
Aroa López-Sánchez ◽  
Laura Tomás-Gallardo ◽  
Eduardo Santero ◽  
Fernando Govantes
Keyword(s):  
Complex Formation ◽  
Promoter Region ◽  
Atp Hydrolysis ◽  
Cyanuric Acid ◽  
Regulatory Protein ◽  
Dna Interaction ◽  
Integration Host Factor ◽  
Open Complex Formation

ABSTRACTPatzTis an internal promoter of theatzRSTUVWoperon that directs the synthesis of AtzT, AtzU, AtzV, and AtzW, components of an ABC-type cyanuric acid transport system. PatzTis σNdependent, activated by the general nitrogen control regulator NtrC with the assistance of protein integration host factor (IHF), and repressed by the LysR-type transcriptional regulator (LTTR) AtzR. We have used a variety ofin vivoandin vitrogene expression and protein-DNA interaction assays to assess the mechanisms underlying AtzR-dependent repression of PatzT. Here, we show that repression only occurs when AtzR and NtrC interact simultaneously with the PatzTpromoter region, indicating that AtzR acts as an antiactivator to antagonize activation by NtrC. Furthermore, repression requires precise rotational orientation of the AtzR and NtrC binding sites, strongly suggesting protein-protein interaction between the two proteins on the promoter region. Further exploration of the antiactivation mechanism showed that although AtzR-dependent repression occurs prior to open complex formation, AtzR does not alter the oligomerization state of NtrC or inhibit NtrC ATPase activity when bound to the PatzTpromoter region. Taken together, these results strongly suggest that PatzT-bound AtzR interacts with NtrC to prevent the coupling of NtrC-mediated ATP hydrolysis with the remodeling of the interactions between E-σNand PatzTthat lead to open complex formation.IMPORTANCEHere, we describe a unique mechanism by which the regulatory protein AtzR prevents the activation of the σN-dependent promoter PatzT. Promoters of this family are always positively regulated, but there are a few examples of overlapping negative regulation. The mechanism described here is highly unconventional and involves an interaction between the repressor and activator proteins to prevent the action of the repressor protein on the RNA polymerase-promoter complex.

scholarly journals Tumor Suppressor HLJ1 Binds and Functionally Alters Nucleophosmin via Activating Enhancer Binding Protein 2  Complex Formation

2010 ◽  
Vol 70 (4) ◽  
pp. 1656-1667 ◽  
Author(s):  
T.-P. Chang ◽  
S.-L. Yu ◽  
S.-Y. Lin ◽  
Y.-J. Hsiao ◽  
G.-C. Chang ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Complex Formation ◽  
Binding Protein ◽  
Enhancer Binding Protein

scholarly journals Isolation and Properties of the Complex between the Enhancer Binding Protein NIFA and the Sensor NIFL

1999 ◽  
Vol 181 (15) ◽  
pp. 4461-4468 ◽  
Author(s):  
Tracy Money ◽  
Tamera Jones ◽  
Ray Dixon ◽  
Sara Austin
Keyword(s):  
Complex Formation ◽  
Azotobacter Vinelandii ◽  
Binding Protein ◽  
Protein Protein Interaction ◽  
Amino Terminal ◽  
Cell Extracts ◽  
Enhancer Binding Protein ◽  
Adenosine Nucleotides

ABSTRACT In Azotobacter vinelandii, activation ofnif gene expression by the transcriptional regulatory enhancer binding protein NIFA is controlled by the sensor protein NIFL in response to changes in levels of oxygen and fixed nitrogen in vivo. NIFL is a novel redox-sensing flavoprotein which is also responsive to adenosine nucleotides in vitro. Inhibition of NIFA activity by NIFL requires stoichiometric amounts of the two proteins, implying that the mechanism of inhibition is by direct protein-protein interaction rather than by catalytic modification of the NIFA protein. The formation of the inhibitory complex between NIFL and NIFA may be regulated by the intracellular ATP/ADP ratio. We show that adenosine nucleotides promote complex formation between purified NIFA and NIFL in vitro, allowing isolation of the NIFL-NIFA complex. The complex can also be isolated from cell extracts containing coexpressed NIFL and NIFA in the presence of MgADP. Removal of the nucleotide causes dissociation of the complex. Experiments with truncated proteins demonstrate that the amino-terminal domain of NIFA and the C-terminal region of NIFL potentiate the ADP-dependent stimulation of NIFL-NIFA complex formation.

scholarly journals Protein-Protein Interactions in the Complex between the Enhancer Binding Protein NIFA and the Sensor NIFL fromAzotobacter vinelandii

2001 ◽  
Vol 183 (4) ◽  
pp. 1359-1368 ◽  
Author(s):  
Tracy Money ◽  
Jason Barrett ◽  
Ray Dixon ◽  
Sara Austin
Keyword(s):  
Complex Formation ◽  
Protein Interactions ◽  
Azotobacter Vinelandii ◽  
Protein Complexes ◽  
Binding Protein ◽  
Limited Proteolysis ◽  
Regulatory System ◽  
Terminal Domain ◽  
Enhancer Binding Protein

ABSTRACT The enhancer binding protein NIFA and the sensor protein NIFL fromAzotobacter vinelandii comprise an atypical two-component regulatory system in which signal transduction occurs via complex formation between the two proteins rather than by the phosphotransfer mechanism, which is characteristic of orthodox systems. The inhibitory activity of NIFL towards NIFA is stimulated by ADP binding to the C-terminal domain of NIFL, which bears significant homology to the histidine protein kinase transmitter domains. Adenosine nucleotides, particularly MgADP, also stimulate complex formation between NIFL and NIFA in vitro, allowing isolation of the complex by cochromatography. Using limited proteolysis of the purified proteins, we show here that changes in protease sensitivity of the Q linker regions of both NIFA and NIFL occurred when the complex was formed in the presence of MgADP. The N-terminal domain of NIFA adjacent to the Q linker was also protected by NIFL. Experiments with truncated versions of NIFA demonstrate that the central domain of NIFA is sufficient to cause protection of the Q linker of NIFL, although in this case, stable protein complexes are not detectable by cochromatography.

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