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 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

scholarly journals Modulation of Mycobacterium tuberculosis DnaAprotein–adenine nucleotide interactions by acidic phospholipids

2002 ◽  
Vol 364 (3) ◽  
pp. 887-888
Author(s):  
K. YAMAMOTO ◽  
S. MUNIRUZZAMAN ◽  
M. RAJAGOPALAN ◽  
M.V.V.S. MADIRAJU
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Adenine Nucleotide ◽  
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.

scholarly journals Sabicea brasiliensis Wernham: Antioxidant Activity, Proliferative Effect and Modulation of Vascular Adenine Nucleotides Metabolism

2020 ◽  
pp. 1-10
Author(s):  
Douglas Souza Oliveira ◽  
Mikaelle Costa Correia ◽  
Bruna Juber de Araújo ◽  
Fernanda Cardoso da Silva ◽  
Paula Marynella Alves Pereira Lima ◽  
...  
Keyword(s):  
Antioxidant Activity ◽  
Cell Viability ◽  
Phenolic Content ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Total Phenolic ◽  
Vascular Cells ◽  
Proliferative Effect ◽  
A7r5 Cells ◽  
Hydrolysis Of

Aims: Study addressed the antioxidant activity (AA) of Sabicea brasiliensis roots crude extract (CE), ethyl acetate (EAF), and hydro-methanolic (HMF) fractions, and its impact on cell viability and adenine nucleotide hydrolysis in vascular A7r5 cells. Materials and Methods: AA of CE, EAF and HMF were determined by the inhibition of the DPPH and ABTS radicals. Total phenolic content was evaluated by Folin-Ciocalteau. Cell viability was determined by MTT assay at different concentrations (62.5; 125; 250 and 500 μg·mL-1) of EAF and HMF after 24, 48 and 72 h. Ectonucleotidase activities were evaluated by colorimetric methods after 48 h EAF or HMF treatment. Results: The highest AA was observed for CE (76%), followed by EAF (46%) and HMF (23%). Phenolic content followed the same pattern. After 48 h, EAF increased A7r5 vascular cells viability by 40%, 40%, 62% and 25% at distinct concentrations, respectively; while HMF augmented it by 50% (500 μg·mL−1). Finally, after 48 h EAF (500 μg·mL−1) decreased about 50% of ATP and ADP metabolism while HMF inhibited 56 and 59% the hydrolysis of NPP substrate (at 125 and 250 μg·mL−1). Conclusion: Study confirmed the high AA of S. brasiliensis, which influences vascular cells proliferation and purines metabolism, pointing to potential cellular pathways that may support the popular use of this plant.

Further studies on the presumed adenine nucleotide storage compound of rat heart

1985 ◽  
Vol 5 (12) ◽  
pp. 1061-1069 ◽  
Author(s):  
Peter S. Fitt ◽  
Borivoj Korecky ◽  
Nishi Sharma
Keyword(s):  
High Resistance ◽  
Alkaline Hydrolysis ◽  
Rat Heart ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Significant Proportion ◽  
Radioactive Substance ◽  
Sudden Rise ◽  
Rat Hearts ◽  
Hydrolysis Of

Further studies on the acid-precipitable radioactive substance formed during perfusion of Langendorff rat hearts with [14C]adenosine have shown that very brief (30 s) ischaemia causes a sudden rise (20–35%) in its level in the tissue which is followed by the steady fall we have previously described. Analysis of the products of alkaline hydrolysis of this compound shows that at least 96% of the radioactivity appears in the form of a mixture of 2′- and 3′-AMP as would be expected for RNA while its relatively high resistance to dilute alkali suggests that it is poly A. Subcellular localization studies indicate that radioactivity enters all compartments of the cell, with maximum label in the nucleus. However, a significant proportion is present in the mitochondria and may be poly A acting as the mitochondrial storage form of adenine nucleotides whose existence we have proposed.

The Platelet of the Newborn Infant – Adenine Nucleotide Metabolism and Release

1980 ◽  
Vol 43 (02) ◽  
pp. 099-103 ◽  
Author(s):  
J M Whaun ◽  
P Lievaart ◽  
Keyword(s):  
Newborn Infant ◽  
Adenine Nucleotide ◽  
Platelet Rich Plasma ◽  
Adenine Nucleotides ◽  
Newborn Infants ◽  
Specific Radioactivity ◽  
Nucleotide Metabolism ◽  
Breakdown Product ◽  
Term Infants ◽  
Adenine Nucleotide Metabolism

SummaryBlood from normal full term infants, mothers and normal adults was collected in citrate. Citrated platelet-rich plasma was prelabelled with 3H-adenine and reacted with release inducers, collagen and adrenaline. Adenine nucleotide metabolism, total adenine nucleotide levels and changes in sizes of these pools were determined in platelets from these three groups of subjects.At rest, the platelet of the newborn infant, compared to that of the mother and normal adult, possessed similar amounts of adenosine triphosphate (ATP), 4.6 ± 0.2 (SD), 5.0 ± 1.1, 4.9 ± 0.6 µmoles ATP/1011 platelets respectively, and adenosine diphosphate (ADP), 2.4 ± 0.7, 2.8 ± 0.6, 3.0 ± 0.3 umoles ADP/1011 platelets respectively. However the marked elevation of specific radioactivity of ADP and ATP in these resting platelets indicated the platelet of the neonate has decreased adenine nucleotide stores.In addition to these decreased stores of adenine nucleotides, infant platelets showed significantly impaired release of ADP and ATP on exposure to collagen. The release of ADP in infants, mothers, and other adults was 0.9 ± 0.5 (SD), 1.5 ± 0.5, 1.5 ± 0.1 umoles/1011 platelets respectively; that of ATP was 0.6 ± 0.3, 1.0 ± 0.1,1.3 ± 0.2 µmoles/1011 platelets respectively. With collagen-induced release, platelets of newborn infants compared to those of other subjects showed only slight increased specific radioactivities of adenine nucleotides over basal levels. The content of metabolic hypoxanthine, a breakdown product of adenine nucleotides, increased in both platelets and plasma in all subjects studied.In contrast, with adrenaline as release inducer, the platelets of the newborn infant showed no adenine nucleotide release, no change in total ATP and level of radioactive hypoxanthine, and minimal change in total ADP. The reason for this decreased adrenaline reactivity of infant platelets compared to reactivity of adult platelets is unknown.Infant platelets may have different membranes, with resulting differences in regulation of cellular processes, or alternatively, may be refractory to catecholamines because of elevated levels of circulating catecholamines in the newborn period.

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 L-Arginine Intake Effect on Adenine Nucleotide Metabolism in Rat Parenchymal and Reproductive Tissues

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
G. Kocic ◽  
J. Nikolic ◽  
T. Jevtovic-Stoimenov ◽  
D. Sokolovic ◽  
H. Kocic ◽  
...  
Keyword(s):  
Adenosine Deaminase ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Tissue Growth ◽  
Nucleotide Metabolism ◽  
Amp Deaminase ◽  
Male Impotence ◽  
Adenine Nucleotide Metabolism ◽  
Adenosine Concentration ◽  
Male Wistar Rats

L-arginine is conditionally essetcial amino acid, required for normal cell growth, protein synthesis, ammonia detoxification, tissue growth and general performance, proposed in the treatment of men sterility and prevention of male impotence. The aim of the present paper was to estimate the activity of the enzymes of adenine nucleotide metabolism:5′-nucleotidase (5′-NU), adenosine deaminase (ADA), AMP deaminase, and xanthine oxidase (XO), during dietary intake of L-arginine for a period of four weeks of male Wistar rats. Adenosine concentration in tissues is maintained by the relative activities of the adenosine-producing enzyme,5′-NU and the adenosine-degrading enzyme-ADA adenosine deaminase. Dietary L-arginine intake directed adenine nucleotide metabolism in liver, kidney, and testis tissue toward the activation of adenosine production, by increased5′-NU activity and decreased ADA activity. Stimulation of adenosine accumulation could be of importance in mediating arginine antiatherosclerotic, vasoactive, immunomodulatory, and antioxidant effects. Assuming that the XO activity reflects the rate of purine catabolism in the cell, while the activity of AMP deaminase is of importance in ATP regeneration, reduced activity of XO, together with the increased AMP-deaminase activity, may suggest that adenine nucleotides are presumably directed to the ATP regenerating process during dietary L-arginine intake.

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
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