scholarly journals Interaction of Trypanosoma cruzi adenylate cyclase with liver regulatory factors

1986 ◽  
Vol 236 (1) ◽  
pp. 185-191 ◽  
Author(s):  
C Eisenschlos ◽  
M M Flawiá ◽  
M Torruella ◽  
H N Torres
Keyword(s):  
Trypanosoma Cruzi ◽  
Adenylate Cyclase ◽  
Rat Liver ◽  
Phospholipid Vesicles ◽  
Guanine Nucleotide ◽  
Catalytic Subunits ◽  
Regulatory Factors ◽  
Liver Membrane ◽  
Catalytically Active ◽  
Liver Membranes

Trypanosoma cruzi adenylate cyclase catalytic subunits may interact with regulatory factors from rat liver membranes, reconstituting heterologous systems which are catalytically active in assay mixtures containing MgATP. The systems show stimulatory responses to glucagon and guanosine 5′-[beta gamma-imido]triphosphate (p[NH]ppG) or fluoride. Reconstitution was obtained by three different methods: fusion of rat liver membranes (pretreated with N-ethylmaleimide) to T. cruzi membranes; interaction of detergent extracts of rat liver membranes with T. cruzi membranes; or interaction of purified preparations of T. cruzi adenylate cyclase and of liver membrane factors in phospholipid vesicles. The liver factors responsible for the guanine nucleotide effect were characterized as the NS protein. Data also indicate that reconstitution requires the presence of a membrane substrate.

Binding of [14,15-3H]14,15-dihydroforskolin to rat liver membranes: comparison with the stimulatory effect of forskolin on adenylate cyclase

1987 ◽  
Vol 65 (5) ◽  
pp. 803-809 ◽  
Author(s):  
Kurt Schmidt ◽  
Hans P. Baer ◽  
Azim Shariff ◽  
William A. Ayer ◽  
Lois Browne
Keyword(s):  
Adenylate Cyclase ◽  
Rat Liver ◽  
Binding Sites ◽  
Binding Capacity ◽  
Protein A ◽  
High Specificity ◽  
Binding Studies ◽  
Liver Membranes ◽  
High Concentrations ◽  
Equilibrium Dissociation

The binding of [14,15-3H]14,15-dihydroforskolin ([3H]DHF) to rat liver membranes has been further characterized and was compared with the stimulatory effect of forskolin on adenylate cyclase. The binding equilibrium dissociation constant (KD) for 14,15-dihydroforskolin obtained in inhibition experiments was 0.6 μM, with a maximal binding capacity (Bmax) of 114 pmol/mg protein. A similar KD value (0.5 μM) was derived from kinetics studies that revealed very rapid association and dissociation reactions. For structure–activity relationship studies several forskolin derivatives were synthesized and tested for their ability to inhibit [3H]DHF binding and increase adenylate cyclase activity. Among the tested compounds, forskolin itself was the most potent agonist (KI = 0.2 μM). Further modification of the molecule in position 7 and (or) 1 decreased or abolished its agonist properties in both adenylate cyclase and binding studies. [3H]DHF binding was not affected by several nucleotides, carbohydrates, lectins, and hormone receptor agonists including isoproterenol, glucagon, and adenosine, but the steroids 17-β-estradiol, progesterone, and testosterone showed slight inhibitory effects at unphysiologically high concentrations, [3H]DHF binding and forskolin-stimulated adenylate cyclase were sensitive to heat and N-ethylmaleimide treatment. Forskolin protected adenylate cyclase against inactivation by heat but not by N-ethylmaleimide. Preincubation of the membrane with trypsin decreased [3H]DHF binding. The results presented in this study demonstrate that the binding sites identified with [3H]DHF have a high specificity for forskolin and provide evidence that these binding sites are involved in the stimulation of adenylate cyclase by forskolin.

Oxytocin and vasopressin analogues capable of competition with vasopressin binding to rat liver membranes

1983 ◽  
Vol 48 (6) ◽  
pp. 1788-1795
Author(s):  
Tomislav Barth ◽  
Bernard Cantau ◽  
Daniel Butlen ◽  
Gilles Guillon ◽  
Serge Jard ◽  
...  
Keyword(s):  
Rat Liver ◽  
Membrane System ◽  
Membrane Receptor ◽  
Peptide Chain ◽  
Liver Membrane ◽  
Liver Membranes ◽  
Vasopressin Analogues ◽  
Good Agreement

A number of vasopressin and oxytocin analogues modified in positions 1, 2, 4, 6 and 9 of the peptide chain were tested regarding their affinity to the rat liver membrane receptor. The affinities were estimated from the ability of the analogues to compete with the binding of tritiated vasopressin to rat liver membranes. In the series of vasopressin agonists, the degree of competition was in good agreement with the corresponding pressor activities. In the case of inhibitors of vasopressin pressor action, binding to the membrane system was also observed.

scholarly journals Proteolysis activates adenylate cyclase in rat liver and AC− lymphoma cell independently of the guanine nucleotide regulatory site

FEBS Letters ◽  
1980 ◽  
Vol 115 (2) ◽  
pp. 260-264 ◽  
Author(s):  
Dominique Stengel ◽  
Pramod M. Lad ◽  
Thor B. Nielsen ◽  
Martin Rodbell ◽  
Jacques Hanoune
Keyword(s):  
Adenylate Cyclase ◽  
Rat Liver ◽  
Lymphoma Cell ◽  
Guanine Nucleotide ◽  
Regulatory Site

Analysis of the lipoprotein binding site of rat liver membranes

1994 ◽  
Vol 72 (3-4) ◽  
pp. 132-142 ◽  
Author(s):  
Lynda Adam ◽  
Louise Brissette
Keyword(s):  
Membrane Proteins ◽  
Rat Liver ◽  
Binding Sites ◽  
Plasma Membranes ◽  
Binding Capacity ◽  
New Information ◽  
Liver Membrane ◽  
Liver Membranes ◽  
Bona Fide ◽  
Lipoprotein Binding

Intermediate density lipoproteins (IDL) were shown to bind to high- and low-affinity binding sites on rat liver membranes. The low-affinity sites were named lipoprotein binding sites (LBS), since they bind all classes of lipoproteins. This study was undertaken to further characterize the interaction of 125I-labelled IDL with the LBS of rat liver membranes to determine the chemical nature of the LBS. We found that the binding of IDL to the LBS is insensitive to EDTA and sensitive to heparin and that it is present on plasma membranes. Also, membranes were pretreated with various enzymes that have an effect on the membrane constituents, and the activity of the LBS on these treated membranes was determined. Our results reveal that the LBS of rat liver membranes is insensitive to heparinase I, chondroitinase ABC, and phospholipase C, while it is partially sensitive to phospholipase A2 and sensitive to proteases and heat. Rat liver membrane proteins were solubilized with Triton X-100, reconstituted in liposomes, and analyzed for their ability to bind lipoproteins. 125I-labelled IDL were shown to bind to high- and low-affinity sites that are similar, in affinity and specificity, to the ones observed with intact rat liver membranes, indicating that a LBS activity is detectable on these liposomes. We found that the binding capacity of low-affinity sites in liposomes containing either no protein or containing proteins solubilized from Escherichia coli membranes is five times weaker than low-affinity sites in liposomes containing liver membrane proteins. Thus, a protein solubilized from rat liver membranes has LBS activity when reconstituted in liposomes. Taken altogether our results provide new information on the binding of IDL to the LBS and indicate that the LBS activity is in part mediated by a protein. Thus, the LBS appears to be a bona fide receptor.Key words: lipoprotein, receptor, binding, metabolism.

scholarly journals The role of a guanine nucleotide-binding protein in the activation of rat liver plasma-membrane adenylate cyclase by forskolin

1983 ◽  
Vol 216 (3) ◽  
pp. 753-759 ◽  
Author(s):  
S K F Wong ◽  
B R Martin
Keyword(s):  
Plasma Membrane ◽  
Adenylate Cyclase ◽  
G Protein ◽  
Rat Liver ◽  
Guanine Nucleotide ◽  
Guanine Nucleotide Binding Protein ◽  
Liver Plasma Membrane ◽  
Guanine Nucleotide Binding ◽  
The Individual

The effects of the photoreactive GTP analogue GTP-gamma-azidoanilide on rat liver plasma-membrane adenylate cyclase are described. U.v. irradiation in the presence of the analogue abolished activation by any effector or combination of effectors that function via the activatory G protein. Partial protection against this inhibition was given by F- and guanosine 5′-[gamma-thio]triphosphate. It is concluded that GTP-gamma-azidoanilide acts by a light-induced covalent reaction with the G protein. In the dark the effects of the analogue were similar to those of GTP. Irradiation in the presence of GTP-gamma-azidoanilide was found to reduce but not to abolish activation of rat liver plasma membrane adenylate cyclase by forskolin. The activation by forskolin and GTP together were greater than the sum of the individual activations. Forskolin doubled adenylate cyclase activity in the presence of glucagon and guanosine 5′-[beta, gamma-imido]triphosphate, which might be expected to activate to the maximum possible extent via the G protein. It is concluded that there are two components to the forskolin activation, a guanine nucleotide-dependent and a guanine nucleotide-independent component.

Non-specific inhibition of some rat liver membrane-bound enzymes by an adenylate cyclase inhibitor, RMI 12330 A

1978 ◽  
Vol 27 (5) ◽  
pp. 641-645 ◽  
Author(s):  
Georges Guellaen ◽  
Jean-Louis Mahu ◽  
Philippe Mavier ◽  
Jacques Hanoune ◽  
Pierre Berthelot
Keyword(s):  
Adenylate Cyclase ◽  
Rat Liver ◽  
Specific Inhibition ◽  
Liver Membrane ◽  
Membrane Bound ◽  
Membrane Bound Enzymes

PACAP and VIP receptors in rat liver membranes

1991 ◽  
Vol 260 (1) ◽  
pp. G97-G102 ◽  
Author(s):  
P. Robberecht ◽  
P. Gourlet ◽  
A. Cauvin ◽  
L. Buscail ◽  
P. De Neef ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Rat Liver ◽  
Plasma Membranes ◽  
Specific Class ◽  
Vip Receptors ◽  
Pituitary Adenylate Cyclase ◽  
Half Maximal Effective Concentration ◽  
Liver Membranes ◽  
Pacap Receptors ◽  
Binding Curves

Pituitary adenylate cyclase activating peptide (PACAP) tested as PACAP-(1-38)NH2 and PACAP-(1-27)NH2 and vasoactive intestinal polypeptide (VIP) were compared for their capacity to discriminate between high- and low-affinity VIP-preferring receptors that coexist in rat liver plasma membranes. This capacity was evaluated by the ability to 1) inhibit 125I-labeled-PACAP-(1-27)NH2, 125I-labeled-VIP, and 125I-labeled-helodermin binding and 2) to activate adenylate cyclase. PACAP-(1-27)NH2 bound specifically and reversibly to three classes of binding sites, as revealed by analysis of binding curves. On high-affinity VIP receptors (tested specifically by [125I]-helodermin binding), PACAP-(1-38)NH2 showed lower affinity than PACAP-(1-27)NH2 and VIP itself. On low-affinity VIP receptors, PACAP-(1-27)NH2 and -(1-38)NH2 showed similar modest affinity that was slightly higher however than that of VIP. For a third specific class of PACAP receptors (20% of PACAP receptors not recognized by VIP), PACAP-(1-38)NH2 showed higher affinity than PACAP-(1-27)NH2. Both PACAPs stimulated rat liver adenylate cyclase with the same low efficacy as VIP but with an affinity even greater [half-maximal effective concentration (EC50) 0.02 nM] than that of VIP (EC50 0.05 nM).

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