A comparison of cobalamin binding by liver and kidney in rat and man

1985 ◽  
Vol 68 (3) ◽  
pp. 357-364 ◽  
Author(s):  
J. S. D. Scott ◽  
E. P. W. Bowman ◽  
W. G. E. Cooksley
Keyword(s):  
Binding Sites ◽  
Plasma Membranes ◽  
Competitive Inhibition ◽  
Rat Kidney ◽  
Binding Constants ◽  
Human Kidney ◽  
Specific Adsorption ◽  
Receptor Protein ◽  
Mean Values ◽  
Liver And Kidney

1. Binding of cobalamin (Cbl) was compared in liver and kidney plasma membranes prepared from rat and human tissues. 2. Single, high-affinity, saturable (200 pmol/l), binding sites for TC II-Cbl were found in all tissues; by contrast no receptors were present for free cobalamin, for which only non-specific adsorption occurred. 3. Binding constants for TC II-CNCbl determined for liver and kidney plasma membranes were of a similar magnitude. Mean values for Ka (litre/nmol) were 16.7 (rat liver), 18.8 (rat kidney), 8.0 (human liver) and 7.5 (human kidney). 4. Results for binding TC II-OHCbl instead of TC II-CNCbl showed no difference in Ka and Bmax. values, although the non-specific adsorption was decreased to a third. 5. Competitive inhibition results showed that the receptors are specific for the TC II molecule and that binding is unaffected either by the cobalamin moiety or by the presence of free cobalamin. Degradation of the receptor protein molecules by trypsin (10 μg/ml) resulted in 90% inhibition of binding. 6. It is concluded that differences between liver and kidney in cobalamin uptake and accumulation cannot be attributed to differences in their TC II receptors.

scholarly journals Uptake of iodinated human kidney α-D-mannosidase by rat liver- Association with membrane elements and stability in vivo and in vitro

1976 ◽  
Vol 159 (3) ◽  
pp. 729-736 ◽  
Author(s):  
D V Marinković ◽  
S L Petrović ◽  
J V Martinović ◽  
J N Marinković
Keyword(s):  
Plasma Membranes ◽  
Specific Radioactivity ◽  
Human Kidney ◽  
Particulate Material ◽  
Enzymic Activity ◽  
Free Form ◽  
Rat Tissues ◽  
Liver And Kidney

1. Human kidney α-D-mannosidase (form A) was labelled with 125I to a specific radio-activity of approx. 2250muCi/mg of protein, essentially without loss of enzymic activity. The enzymic activity and radioactivity of the iodinated material also co-migrated in gel filtration and gel electrophoresis. 2. The binding of 125I-labelled mannosidase in vitro to particulate material in liver and kidney homogenates was of the other of 2 pg/mg of particulate material in liver and kidney homogenates was of the order of 2pg/mg of particulate protein withing 16h at 37 degrees C, and essentially zero in intervals of up to 60 min. The degradation in vitro of labelled exogenous mannosidase was of the order of 10-20pg/ 16th per mg of protein in postnuclear supernatant, and it was saturated entirely within 1h at 37 degrees C. 3. The binding of labelled mannosidase in vivo to particulate elements of liver homogenates 60 min after intravenous injection was at least 10 times higher in terms of specific radioactivity than the highest value attainable in vitro. Virtually all exogenous enzyme bound to liver particulate material could be recovered in macromolecular form after disruption of membranes by detergents. 4. The radioactive enzyme bound to liver particulate material could be detached almost completely by shearing, repeated freezing and thawing, and exposure to strong detergents under conditions that do not eliminate rough-endoplasmic-membrane structure. It could bot be released, however, by high salt concentration (0.5M-KC1) or by exposure to weak detergents such as Tween 80. The particle-bound enzyme should thus be associated with plasma membranes and lysosome-like elements. 5. Of the rat tissues studied, only liver could approach, within 60 min after the injection, the concentration of exogenous mannosidase found in the blood serum. The activity per g tissue weight fell progressively from liver (60% of serum value) to kidney (16% of serum value), lung (8% of serum vlaue), spleen (6% of serum value) and brain (0.9% of serum value). Most of the radioactive enzyme found in tissues other than liver appeared to be present in a free form, whereas in liver more than 50% of the labelled enzyme was associated with membrane elements.

Immunohistochemical mapping of cellular and subcellular distribution of 5-HT1A receptors in rat and human kidneys

1993 ◽  
Vol 264 (1) ◽  
pp. F9-F19 ◽  
Author(s):  
J. R. Raymond ◽  
J. Kim ◽  
R. E. Beach ◽  
C. C. Tisher
Keyword(s):  
Plasma Membranes ◽  
Radioligand Binding ◽  
Rat Kidney ◽  
Lateral Septum ◽  
Receptor Protein ◽  
Binding Studies ◽  
Igg Antibody ◽  
Connecting Tubule ◽  
Specific Localization ◽  
Immunoperoxidase Labeling

Northern blotting studies have demonstrated mRNA for the serotonin 5-HT1A receptor in human neonatal kidney (B. K. Kobilka, T. Frielle, S. Collins, T. Yang-Feng, T. S. Kobilka, U. Francke, R. J. Lefkowitz, and M. G. Caron. Nature Lond. 329: 75-79, 1987). To confirm expression of receptor protein in kidney, we raised antibodies to two peptides derived from the third intracellular loop of the human 5-HT1A receptor. Specific immunoglobulin G (IgG) was purified sequentially on protein A-Sepharose and peptide-Affigel 10 columns. Each IgG was able to: 1) quantitatively immunoprecipitate [3H]8-OH-2-(di-n-propylamino)1,2,3,4-tetrahydronaphthalene ([3H]8-OH-DPAT)-labeled human and rat receptors; 2) immunoblot a new protein in cells transfected with human 5-HT1A receptor DNA; and 3) immunoautoradiographically label areas of rat brain (frontal cortex, hippocampus, and lateral septum) in a highly characteristic pattern similar to that labeled by 125I-Bolton-Hunter-8-methoxy-2-(N-propyl-N-propylamino)Tetralin, a specific 5-HT1A receptor autoradiography ligand. By use of a light microscopic immunoperoxidase labeling technique, incubation of each IgG antibody with sections of rat and human kidney demonstrated an identical pattern of immunoreactivity. Specific labeling of basolateral plasma membranes was detected throughout medullary and cortical thick ascending limbs (TAL), in distal convoluted tubules (DCT), in connecting tubule cells of the connecting tubule, and in principal cells of the initial collecting tubule. There was no labeling in the inner medulla, glomeruli, or blood vessels. The labeling was blocked by preincubation with the corresponding peptide, but not with noncorresponding peptide or carrier protein. There was no labeling with preimmune IgG. Electron microscopic immunoperoxidase labeling confirmed the specific localization of the IgG antibody along the basolateral plasma membrane in all positively staining cells in rat kidney. Radioligand binding studies with the specific 5-HT1A receptor ligand [3H]8-OH-DPAT confirmed the presence of 5-HT1A receptor binding sites in bulk-isolated rat medullary TAL. These studies provide the first evidence that the 5-HT1A receptor is expressed on the basolateral surface of TAL and DCT cells of human and rat kidney. The specific localization to these cells suggests a possible role for the 5-HT1A receptor in the regulation of salt and water transport in mammalian kidney.

Titration Simulations for Understanding of the Binding Equilibrium

2020 ◽  
Author(s):  
Dae Hyup Sohn
Keyword(s):  
Equilibrium Model ◽  
Binding Sites ◽  
Binding Constants ◽  
Reliability Evaluation ◽  
Binding Isotherm ◽  
Guest Chemistry

<p>The reliability evaluation of the predicted binding constants in numerous models is also a challenge for supramolecular host-guest chemistry. Here, I briefly formulate binding isotherm with the derivation of the multivalent equilibrium model for the chemist who wants to determine the binding constants of their compounds. This article gives an in-depth understanding of the stoichiometry of binding equilibrium to take divalent binding equilibria bearing two structurally identical binding sites as an example. The stoichiometry of binding equilibrium is affected by (1) the cooperativity of complex, (2) the concentration of titration media, and (3) the equivalents of guests. The simulations were conducted with simple Python codes.</p>

(Thio)Urea-Functionalized Cavitands as Excellent Receptors for Organic Anions in Polar Media

2004 ◽  
Vol 69 (5) ◽  
pp. 1137-1148 ◽  
Author(s):  
Gennady V. Oshovsky ◽  
Willem Verboom ◽  
David N. Reinhoudt
Keyword(s):  
Binding Sites ◽  
Binding Constants ◽  
Organic Anions ◽  
Isothermal Microcalorimetry ◽  
Polar Media

Ureidocavitand 1 and thioureidocavitand 2 bind in CH3CN organic anions such as acetate, propionate, butyrate, etc. with K values of 2-8 × 105 l mol-1 and 2-9 × 106 l mol-1, respectively, as was determined with isothermal microcalorimetry (ITC). Bringing together four (thio)urea binding sites on a molecular platform gives rise to about 2000 times higher binding constants, compared with those of the corresponding single binding sites. Glucose- and galactose-containing thioureidocavitands 5 and 6 bind acetate in 1:1 CH3CN/water with a K-value of 2.15 × 103 l mol-1.

scholarly journals Competitive Inhibition of Dibasic Amino Acid Transport in Rat Kidney

1962 ◽  
Vol 237 (7) ◽  
pp. 2265-2270
Author(s):  
Leon E. Rosenberg ◽  
Sylvia J. Downing ◽  
Stanton Segal
Keyword(s):  
Amino Acid ◽  
Amino Acid Transport ◽  
Competitive Inhibition ◽  
Rat Kidney ◽  
Acid Transport ◽  
Dibasic Amino Acid

Eliminating Nonspecific Binding Sites for Highly Reliable Immunoassay via Super-resolution Multicolor Fluorescence Colocalization

Nanoscale ◽  
2021 ◽  
Author(s):  
Shenfei Zong ◽  
Yun Liu ◽  
Kuo Yang ◽  
Zhaoyan Yang ◽  
Zhuyuan Wang ◽  
...  
Keyword(s):  
Binding Sites ◽  
Super Resolution ◽  
Specific Adsorption ◽  
Nonspecific Binding ◽  
Nonspecific Adsorption ◽  
Multicolor Fluorescence

Non-specific adsorption in immunoassays has always been a major problem that affects the reliability of assay results. Despite the emergence of various methods which can reduce nonspecific adsorption, a universal...

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