PROGESTERONE-BINDING GLOBULIN AND TESTOSTERONE-BINDING ACTIVITY IN GUINEA PIG SERUM DURING PREGNANCY: RELATIONSHIP TO PROGESTERONE AND OESTROGENS

1976 ◽  
Vol 81 (2) ◽  
pp. 367-378 ◽  
Author(s):  
O. A. Lea ◽  
A. Bessesen ◽  
K. F. Støa
Keyword(s):  
Guinea Pig ◽  
Binding Sites ◽  
Binding Capacity ◽  
Post Partum ◽  
Equilibrium Dialysis ◽  
Binding Activity ◽  
Maximal Level ◽  
Sharp Drop ◽  
Endogenous Oestrogen ◽  
Competitive Protein Binding

ABSTRACT Progesterone levels in serum have been determined throughout pregnancy in guinea-pigs by a competitive protein-binding technique, using pregnant guinea pig plasma protein as binding agent. The concentrations of this protein (progesterone-binding globulin, PBG) as well as the testosteronebinding activity (TBA) have been quantitated by means of equilibrium dialysis. In order to correlate these parameters to the endogenous oestrogen production, the urinary oestrogen excretion was recorded. The concentration of progesterone, PBG and TBA showed a sharp rise on days 14–18 of gestation, reaching a maximal level on days 30–44. The progesterone level thereafter declined significantly towards parturition, followed by a sharp drop post partum. PBG and TBA followed a similar course, the decline towards parturition, however, being non significant. Neither progesterone, PBG nor TBA were significantly correlated to the urinary oestrogen excretion. The concentration of PGB was remarkably high, amounting to 1–2 × 10-5 m during most of the guinea pig pregnancy. This binding capacity generally exceeded the endogenous progesterone concentration by a factor of 20, as calculated on a molar basis. A mean of 64 % of the binding sites was available for testosterone binding, corresponding to a free binding capacity of 400 μg testosterone per 100 ml.

THYROID HORMONE-BINDING INTERACTIONS IN CYTOSOL OF RAT ANTERIOR PITUITARY

1977 ◽  
Vol 85 (2) ◽  
pp. 256-266 ◽  
Author(s):  
Valerie Anne Galton
Keyword(s):  
Thyroid Hormone ◽  
Binding Sites ◽  
Anterior Pituitary ◽  
Binding Capacity ◽  
Relative Effectiveness ◽  
Binding Activity ◽  
Rat Tissues ◽  
Hormone Binding ◽  
Cytosol Fraction ◽  
Binding Interactions

ABSTRACT Thyroxine (T4) and triiodothyronine (T3)-binding interactions in preparations of rat anterior pituitary gland have been studied. T4 is bound primarily to extranuclear binding sites located in the cytosol fraction of the cell. These sites have a medium affinity for T4: Ka = 2.5 × 108 1/mol and a maximum binding capacity (MBC) of 1.15 pmol/mg tissue (wet weight). Binding of T3 to these sites is minimal. The extent of binding of T4 is influenced by the pH of the system and the temperature of incubation. The relative effectiveness of T4 analogues in displacing bound T4 is tetrac > T4 > triac > D-T4 > T3. Similar T4-binding sites are present in other rat tissues, but in all except serum, binding activity is lower than in the pituitary. T4-binding by serum contaminating the pituitary preparations contributed only partially to the total activity observed. Concomitant assessment of T4-binding activity and T4 metabolism in pituitary homogenates prepared at different pH values indicated an inverse relationship between the two processes. The possible role of thyroid hormone binding in cytosol in influencing the intracellular distribution of thyroid hormones is discussed.

CORTICOSTEROID-BINDING ACTIVITY IN SERUM OF MOUSE, RABBIT AND GUINEA PIG DURING PREGNANCY AND LACTATION: POSSIBLE INVOLVEMENT IN THE INITIATION OF LACTATION

1967 ◽  
Vol 55 (1) ◽  
pp. 47-61 ◽  
Author(s):  
Richard R. Gala ◽  
Ulrich Westphal
Keyword(s):  
Guinea Pig ◽  
Sex Difference ◽  
Equilibrium Dialysis ◽  
Late Pregnancy ◽  
Binding Activity ◽  
Multiple Equilibrium ◽  
Endocrine Control ◽  
Corticosteroid Binding Globulin ◽  
Pregnancy And Lactation ◽  
Approximate Amount

ABSTRACT Corticosteroid-binding activity was determined during pregnancy and lactation in the sera of mouse, rabbit and guinea pig. The measurements were made by multiple equilibrium dialysis using 14C labeled corticosteroids at 37° C. There was a marked increase of binding activity in all three species during pregnancy followed by a decline to the levels of nonpregnant animals during lactation. Serum corticosteroid concentrations paralleled but appeared subsequent to changes in corticosteroid-binding activity during pregnancy and lactation. In the mouse, no sex difference in »resting« corticosterone levels was noted, but the corticosteroid-binding activity was almost 3 times higher in the female animal than in the male. This difference was reflected in a greater amount of unbound corticosterone in the male as evidenced by lower thymus weights. Calculation of the approximate amount of unbound corticosteroid in the mouse and rat during pregnancy revealed a decline at mid pregnancy, followed by an increase from late pregnancy through lactation. In the rabbit and guinea pig, unbound corticosteroid values did not follow similar trends. A mechanism is suggested for the initiation of lactation in the rat, involving the corticosteroid-binding globulin and its endocrine control.

Identification of a set of calcium-binding proteins in reticuloplasm, the luminal content of the endoplasmic reticulum

1988 ◽  
Vol 91 (1) ◽  
pp. 61-70 ◽  
Author(s):  
D.R. Macer ◽  
G.L. Koch
Keyword(s):  
Endoplasmic Reticulum ◽  
Calcium Binding ◽  
Binding Proteins ◽  
Binding Capacity ◽  
Equilibrium Dialysis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Calcium Ionophore ◽  
Binding Activity ◽  
Calcium Binding Proteins ◽  
Molecular Weights

A procedure was developed for the isolation of reticuloplasm, the luminal material of the endoplasmic reticulum (ER). A reticuloplasm-rich extract was prepared from a murine plasmacytoma cell line that contains large amounts of ER, by first extracting the cytoplasmic contents using hypotonic lysis to yield ER-rich ‘shells’ followed by mechanical lysis to release the ER contents. The extract contains five major proteins with apparent molecular weights of 100, 75, 60, 58 and 55 (X 10(3] Mr by SDS-polyacrylamide gel electrophoresis. The 100, 75 and 58 (X 10(3] Mr species were identified as the known ER proteins endoplasmin, BiP and PD1, respectively. The ER association of the 60 and 55 (X 10(3] Mr proteins was confirmed by confocal fluorescence microscopy with affinity-purified antibodies. Equilibrium dialysis with isolated reticuloplasm gave a calcium-binding capacity of 300 nmoles calcium per mg protein with half-maximal binding at 3 mM-Ca2+. Purified endoplasmin bound 280 nmoles calcium per mg protein at a calcium concentration of 5 mM-Ca2+. A calcium overlay test revealed that, in addition to endoplasmin, reticuloplasm contained at least three other calcium-binding proteins: i.e. BiP, PDI and the 55 X 10(3) Mr protein, respectively, with endoplasmin and the 55 X 10(3) Mr protein (CRP55) accounting for the major proportion of the calcium-binding activity. Treatment of cells with calcium ionophore led to the specific over-expression of the major calcium-binding reticuloplasmins endoplasmin, BiP and CRP55. These studies show that the lumen of the ER contains a family of proteins with the capacity to bind significant amounts of calcium in the millimolar range and thereby to confer upon the ER the ability to perform a calcium storage function analogous to that of the sarcoplasmic reticulum in muscle cells.

Interference with thyrotropin receptor antibody determination by a spuriously occurring anti-bovine TSH antibody

1986 ◽  
Vol 112 (2) ◽  
pp. 197-203 ◽  
Author(s):  
Makoto Iitaka ◽  
Toshinori Tanikawa ◽  
Yoshiki Sakatsume ◽  
Morifumi Yanagisawa ◽  
Yoshihito Hara ◽  
...  
Keyword(s):  
Binding Sites ◽  
Binding Capacity ◽  
Binding Activity ◽  
Tsh Receptor ◽  
Thyrotropin Receptor ◽  
Pepsin Digestion ◽  
Thyrotropin Receptor Antibody ◽  
Maximum Binding Capacity ◽  
Antibody Determination ◽  
Bovine Tsh

Abstract. Abnormally negative values of thyrotropin binding inhibitor immunoglobulin (TBII) were found in the sera from a patient with Graves' disease. This was due to the presence of potent bovine TSH (bTSH) binding activity in the sera. This activity was demonstrated to be in immunoglobulin G (IgG) with a λ light chain isotype, which was shown to have an affinity for bTSH with a Ka value of 3.5 × 1010 m−1 and a maximum binding capacity of 1.1 × 10−14m/mg IgG. F(ab')2 fragments obtained through pepsin digestion from the patient's IgG retained bTSH binding activity. [125I] bTSH binding to this IgG was inhibited by the TSH receptor. The inhibition was not completely competitive, suggesting the presence of different binding sites for this IgG and the TSH receptor on the TSH molecule. This IgG, however, could not bind labelled human TSH (hTSH). Since neither TSH nor other pituitary derivatives had ever been given to the patient, this bTSH binding activity was considered to be due to a spuriously occurring anti-bTSH antibody.

CHANGES IN CORTISOL BINDING AND METABOLISM DURING NEONATAL DEVELOPMENT IN THE GUINEA-PIG

1980 ◽  
Vol 85 (2) ◽  
pp. 219-227 ◽  
Author(s):  
M. DALLE ◽  
A. EL HANI ◽  
P. DELOST
Keyword(s):  
Guinea Pig ◽  
Clearance Rate ◽  
Guinea Pigs ◽  
Surgical Stress ◽  
Binding Capacity ◽  
Post Partum ◽  
Metabolic Clearance ◽  
Metabolic Clearance Rate ◽  
Male And Female ◽  
Free Cortisol

The metabolic clearance rate and the binding of cortisol in plasma of male and female guinea-pigs were estimated at five stages between birth and weaning. The metabolic clearance rate for cortisol remained low in both sexes until day 10 post partum but increased to adult values by day 20. The level of free cortisol in plasma and the cortisol binding capacity of the plasma were higher at birth than on day 10 post partum; the values found on day 20 were similar to those of the adult guinea-pig. Surgical stress increased levels of cortisol in plasma on day 20 but not at birth.

Free and bound androgens in the female guinea pig during gestation and after parturition and in the offsprings during the perinatal period

1980 ◽  
Vol 95 (1) ◽  
pp. 101-109 ◽  
Author(s):  
N. Rigaudière ◽  
P. Pradier ◽  
P. Delost
Keyword(s):  
Binding Capacity ◽  
Specific Binding ◽  
Equilibrium Dialysis ◽  
Plasma Concentrations ◽  
Perinatal Period ◽  
Maternal Plasma ◽  
Binding Activity ◽  
Low Concentrations ◽  
High Binding Affinity ◽  
Newborn Animals

Abstract. Total amounts of testosterone (T) and dihydrotestosterone (DHT) and their distribution between non-protein-bound (free), albumin-bound and PBG-(progesterone binding globulin)-bound fractions were determined by radioimmunoassay and equilibrium dialysis from plasma samples. The samples were taken from male guinea pigs during the perinatal period and from pregnant females during gestation and after parturition. Plasma proteins of the foetus and newborn animals appeared to have no high binding affinity for androgens. On the other hand albumin having a binding capacity of 56% for testosterone and 75% for DTH irrespective of the total androgen concentration may be considered to be an important low affinity binding protein. Maternal plasma developed a specific binding activity with the appearance of PBG in early pregnancy. Alterations in the binding capacity of PBG for T or DHT paralleled changes in plasma concentrations of both androgens, the highest values being observed on day 48 of pregnancy, with a prompt return to normal after parturition. Irrespective of the total androgen concentration, it was evident that PBG was capable of maintaining the free T and DHT at low concentrations (about 0.20 and 0.17 ng/10 ml, respectively). Besides the specific binding due to PBG a non-specific binding due to albumin was observed. The competition which exists between these two binding systems for T and DHT was evident when the quantity of bound androgens was expressed as a percentage. Neither the sex, nor the number of the foetuses, nor the interaction between the two, was found to have any significant effect in the maternal androgens, whether total or free hormone was considered.

Pharmacological characterization of the binding of [3H]bremazocine in guinea-pig brain: evidence for multiplicity of the κ-opioid receptors

1989 ◽  
Vol 67 (10) ◽  
pp. 1336-1344 ◽  
Author(s):  
Mario Tiberi ◽  
Jacques Magnan
Keyword(s):  
Guinea Pig ◽  
Opioid Receptors ◽  
Binding Sites ◽  
Binding Capacity ◽  
Pharmacological Characterization ◽  
Homogeneous Population ◽  
Site Model ◽  
Pig Brain ◽  
Guinea Pig Brain

In guinea-pig brain, [3H]bremazocine has a binding capacity of 27.2 pmol/g wet tissue, which is statistically different from that of [3H]ethylketazocine (14.7 pmol/g wet tissue) or the sum of the individual binding capacities of μ-, δ-, and κ-selective ligands (15.0 pmol/g wet tissue). Saturation studies of [3H]bremazocine performed in the presence of unlabelled μ-, δ-, and κ-blockers still reveal a homogeneous population of binding sites. [3H]Bremazocine under suppressed conditions displays at these sites a Kd of 2.51 nM with a binding capacity of 9.15 pmol/g wet tissue. We have performed the pharmacological characterization of these additional opioid binding sites. Displacement curves measured with a number of opioid substances were all best fitted to a one-site model. The stereoselectivity of these additional sites was demonstrated by using two groups of stereoisomers. Oripavine and benzomorphan opioids were among the most potent drugs at the [3H]bremazocine sites (μ + δ + κ suppressed). Diprenorphine, bremazocine, cyclazocine, and ethylketazocine displayed apparent affinities constants (1/Ka) of 8.66, 7.57, 21.4, and 38.0 nM, respectively at those sites. The κ-selective drugs U50488, U69593, PD117302, and tifluadom were inhibitors of the binding of [3H]bremazocine at these sites with apparent affinities of 113, 268, 76.9, and 47.9 nM. All μ- or δ-selective drugs tested in this study have caused weak or no inhibition of the binding. Correlation analyses were done between the different affinities measured at the [3H]bremazocine sites (μ + δ + κ suppressed) and those observed at the known μ-, δ-, and κ-sites of the guinea-pig brain. The results were highly and significantly correlated with the κ-binding profile, while no correlation could be established with the μ,- or δ-binding profiles. Moreover, the results reported here do not correspond to a pharmacological profile that belongs to the σ- or ε-receptors. Thus, it is likely that in the guinea-pig brain the [3H]bremazocine sites (μ + δ + κ suppressed) are a subtype of κ-site family, namely κ2-sites. Finally, apparent affinities displayed at those sites are correlated with the biological activity of opioids in the guinea-pig ileum and rat vas deferens, suggesting that those sites could be considered as receptors.Key words: opioid receptors, kappa subtype, guinea-pig brain.

Erythropoietin binding sites in human foetal tissues

1987 ◽  
Vol 116 (4) ◽  
pp. 561-567 ◽  
Author(s):  
F. Pekonen ◽  
K. Rosenlöf ◽  
E.-M. Rutanen ◽  
F. Fyhrquist
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Recombinant Dna ◽  
Binding Capacity ◽  
Specific Binding ◽  
First Trimester ◽  
Binding Activity ◽  
Renin Substrate ◽  
Human Erythropoietin ◽  
Foetal Liver

Abstract. Using 125I labelled recombinant DNA human erythropoietin (EP), we have explored the presence and properties of EP binding sites in foetal human tissues. The EP binding site is present in the foetal liver already during the first trimester of pregnancy. The binding site has an equilibrium association constant of 4.1–6.2 × 109 1/mol and is specific for EP. The cross-reactivities of FSH, TSH, hCG, insulin and renin substrate were less than 0.01%. The EP binding capacity of foetal liver was 5.4–16 fmol/mg membrane protein. In foetal lung tissue, a slight EP binding activity was observed, whereas foetal spleen, muscle, brain, thyroid and placental tissues were virtually devoid of EP binding capacity. The same level of binding was reached at 37°C in 1 h and at 4°C in 24 h. The binding was pH-dependent with maximal specific binding at pH 7.7. SDS-PAGE gel electrophoresis analysis of covalently cross-linked 125I-EP to foetal liver membranes suggested that the EP binding site was composed of two subunits with an apparent mol wt of 41000 and 86000 dalton, respectively.

Binding of zinc to bovine and human milk proteins

1989 ◽  
Vol 56 (2) ◽  
pp. 235-248 ◽  
Author(s):  
Harjinder Singh ◽  
Albert Flynn ◽  
Patrick F. Fox
Keyword(s):  
Ionic Strength ◽  
Binding Sites ◽  
Binding Capacity ◽  
Equilibrium Dialysis ◽  
Whey Proteins ◽  
Milk Proteins ◽  
Association Constants ◽  
Zn Concentration ◽  
Β Lactoglobulin ◽  
Scatchard Plots

SummaryZn binding by whole bovine and human casein and by purified bovine caseins and whey proteins was investigated by equilibrium dialysis. Bovine αs1 casein had the greatest Zn-binding capacity (˜ 11 atoms Zn/mol). Protein aggregation was observed as Zn concentration was increased and- the protein precipitated at a free Zn concentration of 1·7 mM. Zn binding increased with increasing pH in the range 5·4–7·0 and decreased with increasing ionic strength. Competition between Zn and Ca was observed for binding to αs1-casein indicating common binding sites for these two metals. Bovine β-casein bound up to 8 atoms Zn/ mol and precipitated at a free Zn concentration of ˜ 2·5 mM, while K-casein bound 1–2 atoms Zn/mol. Whole bovine and human casein bound 5–8 atoms Zn/mol and precipitated at a free Zn concentration of ˜ 2·0 mM. Scatchard plots for Zn binding to caseins showed upward convexity, possibly due to Zn-induced association of caseins. Apparent average association constants (K¯app) for all caseins were similar (log K¯app 3·0–3·2). Enzymic dephosphorylation of αs1- or whole bovine casein markedly reduced, but did not eliminate, Zn binding. Thus, phosphoserine residues appeared to be the primary Zn-binding sites in caseins. With the exception of bovine serum albumin. which bound over 8 atoms Zn/mol, the bovine whey proteins, β-lactoglobulin, α-lactalbumin and lactotransferrin, had little capacity for Zn binding.

AN IMPROVED METHOD FOR THE ASSAY OF PROGESTERONE BY COMPETITIVE PROTEIN BINDING

1970 ◽  
Vol 63 (2) ◽  
pp. 225-241 ◽  
Author(s):  
B. D. Reeves ◽  
M. L. A. de Souza ◽  
I. E. Thompson ◽  
E. Diczfalusy
Keyword(s):  
Protein Binding ◽  
Binding Sites ◽  
Entire Range ◽  
Improved Method ◽  
Plasma Progesterone ◽  
Extraction And Purification ◽  
Corticosteroid Binding Globulin ◽  
Competitive Protein Binding ◽  
The Individual ◽  
Range Of Values

ABSTRACT An improved method for the assay of plasma progesterone by competitive protein binding is described. The improvement is based upon rigorous control of the variables, the compensation for and standardisation of interfering factors inherent in the method and the use of a human corticosteroid binding globulin, that meets the requirements for sensitivity at levels of 1.0 ng of progesterone and below. The assessment of the reliability of the individual steps in the method as well as that of the complete method is presented. The sensitivity of the method is around 0.2 ng progesterone per ml plasma. Accuracy was measured by adding progesterone in amounts ranging from 0.0 to 1.0 ng to 1.0 ml plasma. There was a linear relationship between the progesterone added and recovered throughout the entire range of values, with a coefficient of correlation (r) of 0.94. Of 52 related steroids tested, none was found which would remain associated with progesterone following extraction and purification and which would also compete with progesterone for binding sites.

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