scholarly journals Identification of serum GH-binding proteins in the goldfish (Carassius auratus) and comparison with mammalian GH-binding proteins

1999 ◽  
Vol 161 (2) ◽  
pp. 255-262 ◽  
Author(s):  
Y Zhang ◽  
TA Marchant
Keyword(s):  
Binding Sites ◽  
Carassius Auratus ◽  
Binding Proteins ◽  
Binding Capacity ◽  
Sea Bream ◽  
Single Class ◽  
Binding Characteristics ◽  
High Affinity ◽  
Goldfish Carassius Auratus ◽  
Reducing Conditions

The present study constitutes the characterization of a specific, high-affinity GH-binding protein (GHBP) in the serum of a teleost, the goldfish (Carassius auratus). GH-binding assay and ligand blotting techniques were employed to identify GHBPs in goldfish serum and hepatocyte culture medium. The binding characteristics and apparent molecular weights (Mr) of goldfish GHBPs were also compared with those of rabbit and rat. LIGAND analysis identified a single class of high-affinity and low-capacity binding sites for iodinated recombinant carp GH (rcGH) in the goldfish serum, with an association constant (Ka) of 20.1x10(9) M-1 and a maximum binding capacity (Bmax) of 161 fmol ml-1 serum. A single class of binding sites for iodinated recombinant sea bream GH and bovine GH (bGH) was also found in goldfish serum, but with a much lower affinity than that of rcGH. The binding affinity for iodinated bGH in rabbit and rat sera was found to be similar to that reported previously. Ligand blotting revealed multiple forms of GHBPs in sera of goldfish, rabbit and rat with Mr ranging from 70 kDa to 400 kDa and 27 kDa to 240 kDa under non-reducing and reducing conditions respectively. A prominent band with Mr of 66 kDa and a minor band with Mr of 27 kDa were observed to occur in sera from all three species under reducing conditions. Iodoacetamide promoted the shedding of three GHBPs with Mr of 25, 40 and 45 kDa from the cultured goldfish hepatocytes. The appearance of all bands was completely inhibited by the presence of excess unlabeled rcGH. Our results provide clear evidence that a GHBP exists in the goldfish and indicate that more information on teleost GHBPs is needed if the physiology of growth in teleosts is to be fully understood.

Evidence for a direct action of GH on haemopoietic cells of a marine fish, the gilthead sea bream (Sparus aurata)

1995 ◽  
Vol 146 (3) ◽  
pp. 459-467 ◽  
Author(s):  
J A Calduch-Giner ◽  
A Sitjà-Bobadilla ◽  
P Álvarez-Pellitero ◽  
J Pérez-Sánchez
Keyword(s):  
Binding Sites ◽  
Sparus Aurata ◽  
Binding Capacity ◽  
Specific Binding ◽  
Head Kidney ◽  
Gilthead Sea Bream ◽  
Sea Bream ◽  
Single Class ◽  
Lower Vertebrate

Abstract Receptors for GH were characterized in the head kidney of gilthead sea bream (Sparus aurata), using radioiodinated and biotinylated ligands. The specific binding of radiolabelled recombinant gilthead sea bream GH (rsbGH) to head kidney membrane preparations was dependent on membrane concentration. Salmon prolactin, salmon gonadotrophin and carp gonadotrophin did not compete for 125I-labelled rsbGH-binding sites. Unlabelled rsbGH competitively displaced 125I-labelled rsbGH bound to head kidney membranes. Scatchard plots were always linear, denoting the presence of a single class of binding sites. The binding affinity (Ka=2·7 × 109 m−1) was equivalent to that found in liver membrane preparations, but the binding capacity (2·5 ±0·30 fmol/mg protein) was 50- to 75-fold lower. To identify the cells which express the GH receptor, head kidney smears were incubated with biotinylated rsbGH, followed by incubation with an avidin–biotin complex conjugated to alkaline phosphatase. The reaction with the new-fuchsin substrate gave a red precipitate, showing a specific and intense labelling in erythroblasts, polychromatophilic erythroblasts and myeloblasts. Noticeable binding was observed in myelocytes and immature granulocytes, tending to disappear at the latter stages of granulocyte maturation. Light but appreciable binding was also observed in monocytes, lymphocytes and acidophilic erythroblasts, whereas it was completely absent in proerythrocytes and erythrocytes. The proliferative action of rsbGH and recombinant human IGF-I on in vitro cultures of head kidney cells was demonstrated by a 5-bromo-2′-deoxy-uridine immunoassay. To our knowledge, this is the first report that provides suitable evidence for a role of GH as a haemopoietic growth and differentiation factor in lower vertebrate species. Journal of Endocrinology (1995) 146, 459–467

Increase in pituitary dopaminergic receptors after monosodium glutamate treatment

1983 ◽  
Vol 245 (3) ◽  
pp. E261-E265
Author(s):  
M. L. Heiman ◽  
N. Ben-Jonathan
Keyword(s):  
Binding Sites ◽  
Anterior Pituitary ◽  
Arcuate Nucleus ◽  
Binding Capacity ◽  
Monosodium Glutamate ◽  
Male Rats ◽  
Single Class ◽  
Binding Characteristics ◽  
Physiological Conditions ◽  
Da Receptors

We investigated whether a decrease in arcuate nucleus dopamine (DA) levels resulting from neonatal treatment with monosodium glutamate (MSG) affects the anterior pituitary DA receptors in adult male rats. MSG treatment resulted in a significant reduction in medial basal hypothalamic (MBH) DA levels, no change in its norepinephrine (NE) and epinephrine (E) concentrations, and a marked increase in circulating prolactin (PRL). Scatchard analyses of DA binding characteristics to anterior pituitary membranes using [3H]spiperone revealed linear plots, suggesting a single class of high-affinity, low-capacity binding sites. The DA binding capacity was significantly higher in MSG-treated rats than in controls with no change in affinity. The data indicate that anterior pituitary DA receptors change in accordance with altered physiological conditions. The increase in the number of DA receptors following destruction of the arcuate nucleus is probably a direct effect of reduced DA levels reaching the anterior pituitary gland.

Human chorionic gonadotropin binding sites in the human endometrium

1993 ◽  
Vol 129 (1) ◽  
pp. 15-19 ◽  
Author(s):  
S Bhattacharya ◽  
J Banerjee ◽  
S Sen ◽  
PR Manna
Keyword(s):  
Metal Ions ◽  
Chorionic Gonadotropin ◽  
Human Chorionic Gonadotropin ◽  
Binding Sites ◽  
Binding Capacity ◽  
Specific Binding ◽  
Human Endometrium ◽  
Binding Characteristics ◽  
High Affinity ◽  
Specific Binding Sites

The existence of high-affinity and low-capacity specific binding sites for luteinizing hormone (LH)/human chorionic gonadotropin (hCG) has been reported in porcine, rabbit and rat uteri. We have identified hCG binding sites in the human endometrium collected from 35–42-year-old ovulatory and anovulatory women. The binding characteristics of hCG to endometrial tissue preparations from ovulatory and anovulatory women showed saturability with high affinity and low capacity. Scatchard plot analysis showed the dissociation constant of specific binding sites in the ovulatory women to be 3.5 × 10−10 mol/l and in anovulatory women to be 3.1 × 10−10 mol/l. The maximum binding capacity varied considerably between ovulatory (3.85 nmol/kg protein) and anovulatory (6.12 nmol/kg protein) endometrium. Among the divalent metal ions tested (Zn2+, Mg2+, Mn2+, Ca2+—4 mol/l), Zn2+ effected a remarkable increase in [125I]hCG binding to the endometrium (p<0.005) whereas Mn2+ showed a marginal increase and other metal ions did not have any effect. Data obtained with human endometrium indicate an influence of the functional state of the ovary on [125I]hCG binding to endometrium.

Fasting alters somatostatin binding to liver membranes of rainbow trout (Oncorhynchus mykiss)

1996 ◽  
Vol 150 (2) ◽  
pp. 179-186 ◽  
Author(s):  
M J Pesek ◽  
M A Sheridan
Keyword(s):  
Rainbow Trout ◽  
Binding Sites ◽  
Binding Capacity ◽  
Specific Binding ◽  
Scatchard Analysis ◽  
Binding Characteristics ◽  
High Affinity ◽  
Rainbow Trout Oncorhynchus Mykiss ◽  
C Terminus ◽  
Liver Membranes

Abstract Somatostatins are a diverse family of peptides that influence various aspects of animal growth, development, and metabolism. Recent work in our laboratory has shown that somatostatins stimulate hepatic lipolysis in rainbow trout. In this study we characterized somatostatin-binding sites in trout hepatic membrane preparations. We also examined changes in binding characteristics brought about by food deprivation. Binding of [Tyr11]-somatostatin-14 (SS-14) was saturable, reversible, and time- and temperature-dependent. Under optimal conditions, [Tyr11]-SS-14 specific binding averaged 5·7 ± 0·3%. While SS-14 and SS-28 (an N-terminally extended form of SS-14 and derived from the same gene as SS-14) displaced [Tyr11]-SS-14 specific binding (ED50 values of approximately 50 nm and 100 nm respectively), salmon SS-25 (containing [Tyr7,Gly10]-SS-14 at its C terminus and presumably derived from a gene different from that giving rise to SS-14/SS-28), except at pharmacological concentrations, did not. Significant specific binding was also detected in brain, esophagus, stomach, upper and lower intestine, pancreas, and adipose tissue. Scatchard analysis suggested the existence of two classes of hepatic somatostatin-binding sites: a high-affinity site with a Kd of 23 nm and Bmax of 1·4 pmol/mg protein and a low-affinity site with a Kd of 379 nm and Bmax of 4·9 pmol/mg protein. Fasting resulted in reduced growth and elevated plasma levels of SS-14 compared with fed animals. SS-14 binding capacity of the high-affinity class in liver membranes isolated from fasted fish increased by 120% over that from fed counter-parts. No difference in Kd for the high-affinity binding class or in either Kd or Bmax of the low-affinity class was noted between fasted and fed animals. These data support the role of the liver as a target of somatostatin and suggest that fasting enhances hepatic sensitivity to SS-14 binding. Journal of Endocrinology (1996) 150, 179–186

scholarly journals Insulin binding characteristics in canine muscle tissue: effects of the estrous cycle phases

2016 ◽  
Vol 36 (8) ◽  
pp. 761-766 ◽  
Author(s):  
Álan G. Pöppl ◽  
Sandra C. Valle ◽  
Félix H.D. González ◽  
Luiz C. Kucharski ◽  
Roselis S.M. da Silva
Keyword(s):  
Insulin Resistance ◽  
Estrous Cycle ◽  
Muscle Tissue ◽  
Binding Sites ◽  
Binding Capacity ◽  
Insulin Binding ◽  
Glycemic Response ◽  
Intravenous Glucose ◽  
Binding Characteristics ◽  
High Affinity

Abstract: Hormonal fluctuations during the different estrous cycle are a well-recognized cause of insulin resistance in bitches, and little is known about insulin receptor binding or post-binding defects associated with insulin resistance in dogs. To evaluate insulin binding characteristics in muscle tissue of bitches during the estrous cycle, 17 owned bitches were used in the study (six in anestrus, five in estrus, and six in diestrus). An intravenous glucose tolerance test (IVGTT) was performed in all patients by means of injection of 1mL/kg of a glucose 50% solution (500mg/kg), with blood sample collection for glucose determination at 0, 3, 5, 7, 15, 30, 45 and 60 minutes after glucose infusion. Muscle samples, taken after spaying surgery, were immediately frozen in liquid nitrogen and then stored at -80 ºC until the membranes were prepared by sequential centrifugation after being homogenized. For binding studies, membranes were incubated in the presence of 20,000cpm of human 125I-insulin and in increasing concentrations of unlabeled human regular insulin for cold saturation. The IVGTT showed no differences among bitches during the estrous cycle regarding baseline glycemia or glycemic response after glucose infusion. Two insulin binding sites - high-affinity and low-affinity ones - were detected by Scatchard analysis, and significant statistical differences were observed in the dissociation constant (Kd1) and maximum binding capacity (Bmax1) of the high-affinity binding sites. The Kd1 for the anestrus group (6.54±2.77nM/mg of protein) was smaller (P<0.001) than for the estrus (28.54±6.94nM/mg of protein) and diestrus (15.56±3.88nM/mg of protein) groups. Bmax1 in the estrus (0.83±0.42nM/mg of protein) and diestrus (1.24±0.24nM/mg of protein) groups were also higher (P<0.001) than the values observed in anestrus (0.35±0.06nM/mg of protein). These results indicate modulation of insulin binding characteristics during different phases of the estrous cycle in dogs, showing that muscle insulin binding affinity for its receptor is reduced during estrus and diestrus. However, this poor hormone-receptor affinity is compensated for by a greater total binding capacity, once there is no difference in patients' glycemic response after an intravenous glucose load.

scholarly journals Diversity in the structures and ligand-binding sites of nematode fatty acid and retinol-binding proteins revealed by Na-FAR-1 from Necator americanus

2015 ◽  
Vol 471 (3) ◽  
pp. 403-414 ◽  
Author(s):  
M. Florencia Rey-Burusco ◽  
Marina Ibáñez-Shimabukuro ◽  
Mads Gabrielsen ◽  
Gisela R. Franchini ◽  
Andrew J. Roe ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Ligand Binding ◽  
Tissue Distribution ◽  
Binding Sites ◽  
Binding Proteins ◽  
Retinol Binding Protein ◽  
Internal Cavity ◽  
Binding Characteristics ◽  
Necator Americanus ◽  
Ligand Binding Sites

Necator americanus fatty acid and retinol-binding protein-1 (Na-FAR-1) is an abundantly expressed FAR from a parasitic hookworm. The present work describes its tissue distribution, structure and ligand-binding characteristics and shows that Na-FAR-1 expands to transport multiple FA molecules in its internal cavity.

scholarly journals Binding of malonyl-CoA to isolated mitochondria. Evidence for high- and low-affinity sites in liver and heart and relationship to inhibition of carnitine palmitoyltransferase activity

1984 ◽  
Vol 222 (3) ◽  
pp. 639-647 ◽  
Author(s):  
M I Bird ◽  
E D Saggerson
Keyword(s):  
Binding Sites ◽  
Maximal Activity ◽  
Liver Mitochondria ◽  
Carnitine Palmitoyltransferase ◽  
Male Rats ◽  
Heart Mitochondria ◽  
Binding Characteristics ◽  
Functional Sites ◽  
High Affinity ◽  
Malonyl Coa

[14C]Malonyl-CoA bound to intact mitochondria isolated from rat liver and heart in a manner consistent with the presence of two independent classes of binding sites in each tissue. The binding characteristics for mitochondria obtained from fed male rats were: for heart, KD(1) = 11-18nM, KD(2) = 30 microM, N1 = 7pmol/mg of protein, N2 = approx. 660pmol/mg of protein; for liver, KD(1) = 0.1 microM, KD(2) = 5.6 microM, N1 = 11pmol/mg of protein, N2 = 165pmol/mg of protein. In the presence of 40 microM-palmitoyl-CoA the characteristics of binding at the high-affinity sites were changed, so that for heart KD(1) = 0.26 microM, with no change in N1 and for liver KD(1) = approx. 2 microM, with N1 increased to approx. 40pmol/mg of protein. Differences between the two tissues in tightness of malonyl-CoA binding at the high-affinity sites explains the considerably greater sensitivity of heart CPT1 (overt form of carnitine palmitoyltransferase) to inhibition by malonyl-CoA [Saggerson & Carpenter, (1981) FEBS Lett. 129, 229-232; McGarry, Mills, Long & Foster (1983) Biochem. J. 214, 21-28]. Starvation (24h) did not change the characteristics of [14C]malonyl-CoA binding to liver mitochondria and did not alter the I50 (concentration giving 50% inhibition) for displacement of [14C]malonyl-CoA by palmitoyl-CoA. Therefore the decreased sensitivity of liver CPT1 to inhibition by malonyl-CoA in starvation [Saggerson & Carpenter (1981) FEBS Lett. 129, 225-228; Bremer (1981) Biochim. Biophys. Acta 665, 628-631] is not explained by differences in malonyl-CoA binding. Percentage occupancy of the high-affinity sites in heart mitochondria by malonyl-CoA correlated closely with percentage inhibition of CPT1 measured under similar conditions. This finding supports the proposal that the high-affinity binding sites are the functional sites mediating inhibition of CPT1 by malonyl-CoA. Similar experiments with liver mitochondria also suggested that the occupancy of high-affinity sites by malonyl-CoA regulates CPT1 activity. 5,5′-Dithiobis-(2-nitrobenzoic acid), which decreased the sensitivity of heart or liver CPT1 to inhibition by malonyl-CoA [Saggerson & Carpenter (1982) FEBS Lett. 137, 124-128], also decreased [14C]malonyl-CoA binding to the high-affinity sites of heart mitochondria. N1 values for [14C]malonyl-CoA binding to high-affinity sites in liver mitochondria were determined in various physiological states which encompassed a 7-fold range of CPT1 maximal activity (fed, starved, pregnant, hypothyroid, foetal). The N1 value did not change in these states.(ABSTRACT TRUNCATED AT 400 WORDS)

Regulation of bombesin receptors on pancreatic acini by cholecystokinin

1989 ◽  
Vol 256 (2) ◽  
pp. G291-G298
Author(s):  
M. Younes ◽  
S. A. Wank ◽  
R. Vinayek ◽  
R. T. Jensen ◽  
J. D. Gardner
Keyword(s):  
Guinea Pig ◽  
Response Curve ◽  
Binding Capacity ◽  
The Other ◽  
Single Class ◽  
Pancreatic Acini ◽  
Cck Receptors ◽  
Cyclic Monophosphate ◽  
High Affinity ◽  
Bombesin Receptors

When guinea pig pancreatic acini are first incubated with the COOH-terminal octapeptide of cholecystokinin (CCK-8), washed, and then reincubated with 125I-[Tyr4]bombesin (125I-[Tyr4]BN) there is a significant decrease in binding of 125I-[Tyr4]BN compared with that observed with pancreatic acini that have been first incubated with no additions. The CCK-8-induced decrease in binding is maximal after 90 min of first incubation is abolished by reducing the temperature of the first incubation from 37 to 4 degrees C or by adding L364,718 to the first incubation and cannot be reproduced by first incubating acini with A23187, 8-bromoadenosine 3',5'-cyclic monophosphate (8Br-cAMP), 8-bromoadenosine 3',5'-cyclic monophosphate (8Br-cGMP), or 12-O-tetradecanoylphorbol 13-acetate. 125I-[Tyr4]BN interacts with a single class of receptors on pancreatic acini, and first incubating acini with CCK-8 decreases the affinity of BN receptors for BN with no change in the maximal binding capacity. CCK-8 does not alter the rate at which bound 125I-[Tyr4]BN dissociates from pancreatic acini; therefore, CCK-8 must alter the rate at which the radiolabeled BN analogue associates with its receptor. Pancreatic acini possess two classes of CCK receptors: one has a high affinity for CCK-8; the other has a low affinity for CCK-8. The dose-response curve for CCK-8-induced inhibition of binding of 125I-[Tyr4]BN appears to to reflect occupation of low-affinity CCK receptors by CCK-8.(ABSTRACT TRUNCATED AT 250 WORDS)

ATP-MgCl2 treatment after trauma-hemorrhage/resuscitation increases hepatocyte P2-purinoceptor binding capacity

1994 ◽  
Vol 266 (6) ◽  
pp. R1810-R1815
Author(s):  
M. S. Mahmoud ◽  
P. Wang ◽  
S. R. Hootman ◽  
S. S. Reich ◽  
I. H. Chaudry
Keyword(s):  
Binding Capacity ◽  
Scatchard Analysis ◽  
Binding Characteristics ◽  
Beneficial Effects ◽  
High Affinity ◽  
Receptor Component ◽  
Receptor Dynamics ◽  
Maximum Binding Capacity ◽  
High Affinity Receptor ◽  
Affinity Receptor

Although our studies indicate that P2-purinoceptor binding capacity decreases after hemorrhage and resuscitation, it is not known whether ATP-MgCl2 administration after hemorrhage has any beneficial effects on the receptor dynamics. To study this, we performed laparotomy (i.e., trauma induced) on rats and bled them to and maintained them at a mean arterial pressure of 40 mmHg until 40% of maximum bleedout volume was returned in the form of Ringer lactate (RL). The animals were then resuscitated with 3 times the volume of maximum bleedout with RL over 45 min followed by 2 times RL along with ATP-MgCl2 (50 mumol/kg body wt) over 95 min. Hepatocytes were isolated at 4, 17, and 27 h after resuscitation. P2-purinoceptor binding characteristics were determined by using [alpha-35S]ATP. Scatchard analysis revealed high-affinity and low-affinity receptor components in the hepatocytes isolated from sham-operated or hemorrhaged animals with or without ATP-MgCl2 infusion. ATP-MgCl2 ameliorated and subsequently restored the decreased maximum binding capacity (Bmax) of the high-affinity receptor component and significantly improved Bmax of the low-affinity receptor component. ATP-MgCl2 administration also produced a progressive enhancement in the affinity of the low-affinity receptor component. Thus the beneficial effects of ATP-MgCl2 observed after trauma-hemorrhage and resuscitation may be, in part, due to the restoration of P2-purinoceptor binding capacity and the enhancement of the receptor affinity.

CORTICOSTERONE BINDING BY MOUSE SERA DURING FOETAL AND POST-NATAL DEVELOPMENT

1977 ◽  
Vol 84 (1) ◽  
pp. 177-190 ◽  
Author(s):  
Lia Savu ◽  
Emmanuel Nunez ◽  
Max-Fernand Jayle
Keyword(s):  
Amniotic Fluid ◽  
Binding Sites ◽  
Binding Proteins ◽  
Serum Proteins ◽  
Equilibrium Dialysis ◽  
Normal Adult ◽  
Scatchard Analysis ◽  
Mammalian Development ◽  
Single Class ◽  
Mean Values

ABSTRACT The binding properties of corticosterone binding globulin (CBG) of mouse sera have been studied by equilibrium dialysis and electrophoretic techniques, at different stages of foetal and post-natal development. Scatchard analysis has demonstrated in all cases a single class of high affinity saturable binding sites for corticosterone. Remarkable increases of the binding capacities were observed in the foetal and pregnant sera, as compared to normal adult and immature levels. The mean values of n1M1 × g−1 of serum proteins (concentration of binding sites, n1 × moles of binding proteins M1) were 21 10−8 in 14–19 day pregnant females, 17 10−8 in the amniotic fluid, 4.2 10−8 in 14–19 day embryos, and only 0.8 10−8 in the normal adult female. Neonatal mice, aged 0–6 days exhibited no CBG activities. The association constants showed values of 2.5–4.1 108 m−1 when measured with foetal sera, and of 1.2–2.1 108 m−1 with pregnant or control adult sera and with the amniotic fluid, at 25°C. Comparative electrophoretic, thermal denaturation and competition studies with foetal and pregnant plasma CBG's are also reported. The results are discussed in relation to the origin of CBG in the foetal serum, and also with respect to similar studies in the rat, guinea pig and man. The possible biological implications of serum steroid binding proteins in mammalian development are briefly outlined.

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