scholarly journals Molecular Mechanism for Functional Interaction between DnaA Protein and Acidic Phospholipids

2000 ◽  
Vol 276 (10) ◽  
pp. 7450-7456 ◽  
Author(s):  
Masaki Makise ◽  
Shinji Mima ◽  
Tomofusa Tsuchiya ◽  
Tohru Mizushima
Keyword(s):  
Molecular Mechanism ◽  
Functional Interaction ◽  
Dnaa Protein ◽  
Acidic Phospholipids

scholarly journals Involvement of Arg-328, Arg-334 and Arg-342 of DnaA protein in the functional interaction with acidic phospholipids

1999 ◽  
Vol 340 (2) ◽  
pp. 433 ◽  
Author(s):  
Yoshihiro YAMAGUCHI ◽  
Masakazu HASE ◽  
Masaki MAKISE ◽  
Shinji MIMA ◽  
Takeshi YOSHIMI ◽  
...  
Keyword(s):  
Functional Interaction ◽  
Dnaa Protein ◽  
Acidic Phospholipids

scholarly journals Identification of Amino Acids Involved in the Functional Interaction between DnaA Protein and Acidic Phospholipids

2000 ◽  
Vol 275 (6) ◽  
pp. 4513-4518 ◽  
Author(s):  
Masaki Makise ◽  
Shinji Mima ◽  
Tomofusa Tsuchiya ◽  
Tohru Mizushima
Keyword(s):  
Amino Acids ◽  
Functional Interaction ◽  
Dnaa Protein ◽  
Acidic Phospholipids

scholarly journals Involvement of Arg-328, Arg-334 and Arg-342 of DnaA protein in the functional interaction with acidic phospholipids

1999 ◽  
Vol 340 (2) ◽  
pp. 433-438 ◽  
Author(s):  
Yoshihiro YAMAGUCHI ◽  
Masakazu HASE ◽  
Masaki MAKISE ◽  
Shinji MIMA ◽  
Takeshi YOSHIMI ◽  
...  
Keyword(s):  
Amino Acids ◽  
Atp Binding ◽  
Amphipathic Helix ◽  
Wild Type ◽  
Functional Interaction ◽  
Basic Amino Acids ◽  
Dnaa Protein ◽  
Acidic Amino Acids ◽  
Wild Type Protein ◽  
Acidic Phospholipids

We reported previously that three basic amino acids (Arg-360, Arg-364 and Lys-372) of DnaA protein are essential for its functional interaction with cardiolipin. In this study, we examined the effect of mutation of some basic amino acids in a potential amphipathic helix (from Lys-327 to Ile-345) of DnaA protein on this interaction. ATP binding to the mutant DnaA protein, in which Arg-328, Arg-334 and Arg-342 were changed to acidic amino acids, was less inhibited by cardiolipin than that of the wild-type protein, as was the case for mutant DnaA protein with mutations of Arg-360, Arg-364 and Lys-372. A mutant DnaA protein with mutations of all six basic amino acids showed the most resistance to the inhibition of ATP binding by cardiolipin. These results suggest that Arg-328, Arg-334 and Arg-342, like Arg-360, Arg-364 and Lys-372, are also involved in the functional interaction between DnaA protein and acidic phospholipids.

scholarly journals DnaA, the Initiator of Escherichia coliChromosomal Replication, Is Located at the Cell Membrane

2000 ◽  
Vol 182 (9) ◽  
pp. 2604-2610 ◽  
Author(s):  
Gillian Newman ◽  
Elliott Crooke
Keyword(s):  
Cell Membrane ◽  
Spatial Organization ◽  
Active Form ◽  
Chromosomal Replication ◽  
E Coli ◽  
Dnaa Protein ◽  
Section Electron ◽  
Acidic Phospholipids

ABSTRACT Given the lack of a nucleus in prokaryotic cells, the significance of spatial organization in bacterial chromosome replication is only beginning to be fully appreciated. DnaA protein, the initiator of chromosomal replication in Escherichia coli, is purified as a soluble protein, and in vitro it efficiently initiates replication of minichromosomes in membrane-free DNA synthesis reactions. However, its conversion from a replicatively inactive to an active form in vitro occurs through its association with acidic phospholipids in a lipid bilayer. To determine whether the in situ residence of DnaA protein is cytoplasmic, membrane associated, or both, we examined the cellular location of DnaA using immunogold cryothin-section electron microscopy and immunofluorescence. Both of these methods revealed that DnaA is localized at the cell membrane, further suggesting that initiation of chromosomal replication in E. coli is a membrane-affiliated event.

scholarly journals Modulation of Mycobacterium tuberculosis DnaA protein‒adenine-nucleotide interactions by acidic phospholipids

2002 ◽  
Vol 363 (2) ◽  
pp. 305 ◽  
Author(s):  
Kohji YAMAMOTO ◽  
Syed MUNIRUZZAMAN ◽  
Malini RAJAGOPALAN ◽  
Murty V. V. S. MADIRAJU
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Adenine Nucleotide ◽  
Dnaa Protein ◽  
Nucleotide Interactions ◽  
Acidic Phospholipids

Biochemical analysis of DnaA protein with mutations in both Arg328 and Lys372

2002 ◽  
Vol 362 (2) ◽  
pp. 453-458 ◽  
Author(s):  
Masaki MAKISE ◽  
Shinji MIMA ◽  
Motohiro KOTERASAWA ◽  
Tomofusa TSUCHIYA ◽  
Tohru MIZUSHIMA
Keyword(s):  
Biochemical Analysis ◽  
Adenine Nucleotides ◽  
Order Structure ◽  
Chromosomal Dna ◽  
Biochemical Characteristics ◽  
Dnaa Protein ◽  
Acidic Phospholipids ◽  
Mutant Forms

The DnaA protein is the initiator of chromosomal DNA replication in Escherichia coli. Acidic phospholipids decrease its affinity for adenine nucleotides, and re-activate the ADP-bound form to the ATP-bound form. We have previously reported that two mutant forms, DnaAR328E and DnaAK372E, have decreased affinity for cardiolipin (CL). In the present study, we constructed a mutant DnaA protein, DnaA435, with both R328E and K372E, and compared its biochemical characteristics with those of DnaAR328E and DnaAK372E. DnaA435 could bind to oriC DNA, but did not bind ATP or ADP. In DnaA435, compared with DnaAR328E and DnaAK372E, CL caused less inhibition of oriC DNA binding, suggesting that amino acids R328 and K372 are involved in the interaction of DnaA with acidic phospholipids. DnaA435 could initiate DNA synthesis on oriC both in vivo and in vitro. Based on these results, we propose that ATP activates DnaA protein by changing its higher order structure around R328 and K372.

Modulation of Mycobacterium tuberculosis DnaA protein–adenine-nucleotide interactions by acidic phospholipids

2002 ◽  
Vol 363 (2) ◽  
pp. 305-311
Author(s):  
Kohji YAMAMOTO ◽  
Syed MUNIRUZZAMAN ◽  
Malini RAJAGOPALAN ◽  
Murty V.V.S. MADIRAJU
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Membrane Phospholipids ◽  
Membrane Bilayer ◽  
Dnaa Protein ◽  
Head Groups ◽  
Nucleotide Interactions ◽  
Acidic Phospholipids ◽  
Hydrolysis Of

The biochemical aspects of the initiation of DNA replication in Mycobacterium tuberculosis are unknown. To understand this process, we overproduced, purified and characterized the recombinant M. tuberculosis DnaA protein. The M. tuberculosis DnaA protein binds the origin of replication (oriC), ATP and ADP, and exhibited weak ATPase activity. ADP, after hydrolysis of ATP, remained strongly associated with DnaA and the exchange of ATP for bound ADP was weak. Vesicles prepared from acidic phospholipids, such as phosphatidylinositol, cardiolipin and phosphatidylglycerol, promoted dissociation of both ADP and ATP, whereas the neutral phospholipid phosphatidylethanolamine did not. The phospholipid-mediated dissociation of ATP was decreased in the presence of the M. tuberculosis oriC, whereas dissociation of ADP was stimulated in the presence of oriC. Acidic phospholipids in micelles, however, were not efficient in dissociating bound nucleotides from DnaA. Together, these results suggest that both polar head groups and membrane bilayer structure play an important role in M. tuberculosis DnaA—adenine-nucleotide interactions. We suggest that initiation of M. tuberculosis oriC involves intimate interactions between DnaA, adenine nucleotides and membrane phospholipids, and the latter helps to ensure that only the ATP form of the DnaA protein interacts continuously with oriC.

scholarly journals Influence of Cluster Formation of Acidic Phospholipids on Decrease in the Affinity for ATP of DnaA Protein

1996 ◽  
Vol 271 (7) ◽  
pp. 3633-3638 ◽  
Author(s):  
Tohru Mizushima ◽  
Yuichi Ishikawa ◽  
Eishi Obana ◽  
Masakazu Hase ◽  
Toshio Kubota ◽  
...  
Keyword(s):  
Cluster Formation ◽  
Dnaa Protein ◽  
Acidic Phospholipids

scholarly journals Functional interaction of c-Ets-1 and GHF-1/Pit-1 mediates Ras activation of pituitary-specific gene expression: mapping of the essential c-Ets-1 domain.

1995 ◽  
Vol 15 (5) ◽  
pp. 2849-2857 ◽  
Author(s):  
A P Bradford ◽  
K E Conrad ◽  
C Wasylyk ◽  
B Wasylyk ◽  
A Gutierrez-Hartmann
Keyword(s):  
Transcription Factor ◽  
Molecular Mechanism ◽  
Specific Gene ◽  
Ras Signaling ◽  
Cell Type ◽  
Functional Interaction ◽  
Amino Terminal ◽  
Ras Activation ◽  
A Cell ◽  
Cell Type Specific

The mechanism by which activation of common signal transduction pathways can elicit cell-specific responses remains an important question in biology. To elucidate the molecular mechanism by which the Ras signaling pathway activates a cell-type-specific gene, we have used the pituitary-specific rat prolactin (rPRL) promoter as a target of oncogenic Ras and Raf in GH4 rat pituitary cells. Here we show that expression of either c-Ets-1 or the POU homeo-domain transcription factor GHF-1/Pit-1 enhance the Ras/Raf activation of the rPRL promoter and that coexpression of the two transcription factors results in an even greater synergistic Ras response. By contrast, the related GHF-1-dependent rat growth hormone promoter fails to respond to Ras or Raf, indicating that GHF-1 alone is insufficient to mediate the Ras/Raf effect. Using amino-terminal truncations of c-Ets-1, we have mapped the c-Ets-1 region required to mediate the optimal Ras response to a 40-amino-acid segment which contains a putative mitogen-activated protein kinase site. Finally, dominant-negative Ets and GHF constructs block Ras activation of the rPRL promoter, and each blocks the synergistic activation mediated by the other partner protein, further corroborating that a functional interaction between c-Ets-1 and GHF-1 is required for an optimal Ras response. Thus, the functional interaction of a pituitary-specific transcription factor, GHF-1, with a widely expressed nuclear proto-oncogene product, c-Ets-1, provides one important molecular mechanism by which the general Ras signaling cascade can be interpreted in a cell-type-specific manner.

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