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The presence in serum of proteins which are immunologically cross-reactive with Tamm-Horsfall glycoprotein.

ABSTRACT: Affinity chromatography, with rabbit anti-(human Tamm-Horsfall glycoprotein) IgG, was applied to the isolation from normal human serum of protein, which is immunologically cross-reactive with the urinary glycoprotein. The antigen-antibody complex was dissociated with the use of sodium thiocyanate solution, a medium which fails to dissociate urinary Tamm-Horsfall glycoprotein-antigen complex. The cross-reactive serum proteins were isolated in amounts of 19-24 mg/l of serum. They have apparent molecular weights, assessed by disc-gel electrophoresis in the presence of sodium dodecyl sulphate, of 125 000, 84 000 and 74 000 respectively, with mobilities differing from that of urinary Tamm-Horsfall glycoprotein. They have a much lower immunoreactivity towards the antibody than does the urinary glycoprotein. Tamm-Horsfall glycoprotein could not be demonstrated in normal serum by the techniques used. The implications of these findings are discussed in terms of pathology involving Tamm-Horsfall glycoprotein.

SUBMITTER: Lynn KL

PROVIDER: S-EPMC1162780 | biostudies-other | 1981 Feb

REPOSITORIES: biostudies-other

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Subunits of Tamm-Horsfall glycoprotein.

Project description:1. The effect of 6m-guanidine hydrochloride on the urinary glycoprotein described by Tamm & Horsfall (1952) is to produce a homogeneous subunit of molecular weight approx. 100000. 2. Complete reduction of the disulphide bonds of this subunit does not decrease the molecular weight, suggesting that all disulphide bonds are intrachain. 3. Comparison of sedimentation and viscometric behaviour of unreduced and reduced material in 6m-guanidine hydrochloride is consistent with reduction causing an opening-up of intrachain disulphide bonds to give a more asymmetric molecule.

| S-EPMC1185500 | biostudies-other

Some factors affecting the production, by cultured baby-hamster kidney cells, of BHK glycoprotein I which cross-reacts immunologically with Tamm-Horsfall glycoprotein.

Project description:Cultured baby-hamster kidney cells (BHK-21/C13), which are adapted to grow in suspension (strain 2P), roduce a glycoprotein, termed BHK glycoprotein I, which cross-reacts immunologically with hamster urinary (Tamm-Horsfall glycoprotein. BHK glycoprotein I was isolated in an electrophoretically (sodium dodecyl sulphate/polyacrylamide gel) homogeneous form by application of affinity chromatography to the medium in which cells had been cultured. Insolubilized anti-(Tamm-Horsfall glycoprotein immunoglobulin G) was used as the adsorbent. The amount of BHK glycoprotein I associated with the cultured cells was found by both radioimmunoassay and immunofluorescence to be related to the amount of Ca2+ in the medium and to the particular stage of the cell cycle. 5'-Nucleotidase was also shed by the cells into the culture medium in amounts related to the stage of the cell cycle. The turnover of hamster Tamm-Horsfall glycoprotein in vivo appeared to be considerably more rapid than can be accounted for by cell turnover. Hamster Tamm-Horsfall glycoprotein was shown to be ineffective in inhibiting agglutination of chicken erythrocytes caused by influenza virus.

| S-EPMC1164756 | biostudies-other

The development of a radioimmunoassay for Tamm--Horsfall glycoprotein in serum.

Project description:The Tamm-Horsfall glycoprotein prepared by salt precipitation from urine was found to comprise a heterogeneous collection of aggregates. These could be disaggregated with 8m-urea, following which chromatography on a column of Bio-Gel A.15m yielded a homogeneous glycoprotein of mol.wt. 73000 together with several unidentified impurities. Gel filtration of normal plasma showed the glycoprotein to exist predominantly in a form that is eluted identically with the purified preparation. In one case, material of higher molecular weight was also detected. The purified glycoprotein was used to develop a rapid specific radioimmunoassay for its measurement in human serum or plasma by the use of the Tamm-Horsfall glycoprotein, labelled with (125)I by the chloramine-t method as the tracer, an antiserum raised in rabbits, and separation of the bound and free fractions by a second antibody covalently linked to magnetizable particles. Parallelism was demonstrated between the standard preparation and samples. Recovery of added standard to serum varied between 99 and 109%. Total assay time was less than 4h with an intra-assay and inter-assay coefficient of variation of less than 10%. There were no significant differences in the ranges covered with regard to either age or sex, and no circadian rhythm was observed in normal subjects. A physiological range of 70-540ng/ml was established based on serum samples from 95 subjects with normal renal function, as defined by their serum creatinine and urea concentrations. No Tamm-Horsfall glycoprotein was detected in the serum of six anephric patients.

| S-EPMC1161445 | biostudies-other

Tamm-Horsfall urinary glycoprotein. The subunit structure.

Project description:1. Subunit molecular weights of 76000-82000 were obtained for native and alkylated Tamm-Horsfall glycoprotein by gel filtration on Sephadex G-200 in the presence of sodium dodecyl sulphate. 2. A further estimate of the subunit molecular weight of 79000+/-4000 was obtained by disc gel electrophoresis in sodium dodecyl sulphate. 3. A minimum value of the chemical molecular weight of 79000+/-6000 was obtained from the number of N-terminal amino acids released by cyanogen bromide cleavage of the glycoprotein. 4. Similar values were obtained for the subunit molecular weight of Tamm-Horsfall glycoprotein from patients with cystic fibrosis. 5. On ultracentrifugation both in 1.0% sodium dodecyl sulphate and in 70% formic acid, Tamm-Horsfall glycoprotein sedimented as a single component, slightly faster than serum albumin. 6. On reduction of the disulphide bonds the same subunit molecular weight was obtained, which suggested that these bonds are intrachain.

| S-EPMC1179614 | biostudies-other

Tamm-Horsfall urinary glycoprotein. The chemical composition.

Project description:1. A revised amino acid and carbohydrate composition of human Tamm-Horsfall glycoprotein is presented. 2. No significant differences were obtained in the amino acid composition of Tamm-Horsfall glycoprotein isolated from patients with cystic fibrosis. 3. The glycoprotein was shown to possess a high half-cystine content of 1 per 11-12 amino acid residues, which has been confirmed by performic acid oxidation and S-alkylation with iodoacetate and iodoacetamide. No thiol groups were detected in the glycoprotein. 4. Treatment of the glycoprotein with 0.5m-sodium hydroxide at 4 degrees C for 2 days did not release heterosaccharide material, which suggests that the predominant carbohydrate-protein linkages present are not of the O-glycosidic type. 5. No N-terminal amino acid was detected in the glycoprotein.

| S-EPMC1179613 | biostudies-other

Tamm-Horsfall glycoprotein engages human Siglec-9 to modulate neutrophil activation in the urinary tract.

Project description:Urinary tract infections are a major problem in human medicine for which better understanding of native immune defenses may reveal new pathways for therapeutic intervention. Tamm-Horsfall glycoprotein (THP), the most abundant urinary protein, interacts with bacteria including uropathogenic Escherichia coli (UPEC) as well host immune cells. In addition to its well-studied functions to antagonize bacterial colonization, we hypothesize that THP serves a critical host defense function through innate immune modulation. Using isolated human neutrophils, we found that THP binds neutrophils and that this interaction reduces reactive oxygen species generation, chemotaxis and killing of UPEC. We discovered that THP engages the inhibitory neutrophil receptor sialic acid-binding Ig-like lectin-9 (Siglec-9), and mouse functional ortholog Siglec-E, in a manner dependent on sialic acid on its N-glycan moieties. THP-null mice have significantly more neutrophils present in the urine compared with wild-type mice, both with and without the presence of inflammatory stimuli. These data support THP as an important negative regulator of neutrophil activation in the urinary tract, with dual functions to counteract bacterial colonization and suppress excessive inflammation within the urinary tract.

| S-EPMC5698129 | biostudies-literature

Tamm-Horsfall glycoprotein interacts with renal outer medullary potassium channel ROMK2 and regulates its function.

Project description:Tamm-Horsfall glycoprotein (THGP) or Uromodulin is a membrane protein exclusively expressed along the thick ascending limb (TAL) and early distal convoluted tubule (DCT) of the nephron. Mutations in the THGP encoding gene result in Familial Juvenile Hyperuricemic Nephropathy (FJHN), Medullary Cystic Kidney Disease type 2 (MCKD-2), and Glomerulocystic Kidney Disease (GCKD). The physicochemical and biological properties of THGP have been studied extensively, but its physiological function in the TAL remains obscure. We performed yeast two-hybrid screening employing a human kidney cDNA library and identified THGP as a potential interaction partner of the renal outer medullary potassium channel (ROMK2), a key player in the process of salt reabsorption along the TAL. Functional analysis by electrophysiological techniques in Xenopus oocytes showed a strong increase in ROMK current amplitudes when co-expressed with THGP. The effect of THGP was specific for ROMK2 and did not influence current amplitudes upon co-expression with Kir2.x, inward rectifier potassium channels related to ROMK. Single channel conductance and open probability of ROMK2 were not altered by co-expression of THGP, which instead increased surface expression of ROMK2 as determined by patch clamp analysis and luminometric surface quantification, respectively. Despite preserved interaction with ROMK2, disease-causing THGP mutants failed to increase its current amplitude and surface expression. THGP(-/-) mice exhibited increased ROMK accumulation in intracellular vesicular compartments when compared with WT animals. Therefore, THGP modulation of ROMK function confers a new role of THGP on renal ion transport and may contribute to salt wasting observed in FJHN/MCKD-2/GCKD patients.

| S-EPMC3023518 | biostudies-literature

Properties of a glycopeptide isolated from human Tamm-Horsfall glycoprotein. Interaction with leucoagglutinin and anti-(human Tamm-Horsfall glycoprotein) antibodies.

Project description:A sialylated glycopeptide isolated after Pronase digestion of human Tamm-Horsfall glycoprotein behaves as a powerful monovalent hapten in the precipitin reaction between human Tamm-Horsfall glycoprotein and leucoagglutinin, but fails to inhibit the interaction of the glycoprotein with rabbit anti-(human Tamm-Horsfall glycoprotein) antibodies. The glycopeptide is much less active than the intact glycoprotein as an inhibitor of lymphocyte transformation induced by leucoagglutinin.

| S-EPMC1161820 | biostudies-other

Tamm-Horsfall protein and urinary exosome isolation.

Project description:Urinary exosomes have been proposed as starting material for discovery of protein biomarkers of kidney disease. Current protocols for their isolation use a two-step differential centrifugation process. Due to their low density, exosomes are expected to remain in the low-speed (17,000 x g) supernatant and to sediment only when the sample is spun at high speed (200,000 x g). Analysis using western blot and electron microscopy found that urinary exosomes are also present in the low-speed pellet entrapped by polymeric Tamm-Horsfall protein, thus diminishing the procedure's reproducibility. Here we show that addition of dithiothreitol to the low-speed pellet disrupted the polymeric network, presumably by reduction of disulfide bonds linking the monomers. This modification shifted the exosomal proteins from the low- to the high-speed pellet. Also, by shifting the Tamm-Horsfall protein to the high-speed pellet, the use of dithiothreitol makes it feasible to use Tamm-Horsfall protein to normalize excretion rates of exosomal proteins in spot urines. We tested this by western blot, and found that there was a high degree of correlation between exosomal proteins and Tamm-Horsfall protein in the high-speed pellet. Since the yield of exosomes by differential centrifugation can be increased by chemical reduction, Tamm-Horsfall protein may be a suitable normalizing variable for urinary exosome studies when quantitative urine collections are not practical.

| S-EPMC3398710 | biostudies-literature

The turnover rate of rabbit urinary Tamm-Horsfall glycoprotein.

Project description:1. The turnover rate of urinary Tamm-Horsfall glycoprotein in rabbits was determined by two different methods. The first involved measurement of the pool size of the glycoprotein in rabbit kidney and the daily urinary excretion rate by a radioimmunoassay from which the turnover rate was calculated. 2. The second method made use of the incorporation in vivo of Na(2) (14)CO(3) and sodium [(14)C]acetate. After a single intramuscular injection of one of these compounds, urine collections were made every 24h and the glycoprotein was isolated and its specific radioactivity was determined. 3. Incorporation of the label into urinary HCO(3) (-), urea and plasma fibrinogen was also examined. The specific radio-activities of the O-acetyl, sialic acid, aspartic acid and glutamic acid residues isolated from the Tamm-Horsfall glycoprotein were compared and their half-lives were compared with that of the intact glycoprotein. The two methods gave results in quite close agreement and indicated a half-life for the glycoprotein of approx. 9h. 4. An attempt was made to localize the glycoprotein within the kidney and within the cell. It is present throughout the kidney, but was not detected in the brush-border fraction isolated from the proximal tubules. From differential cell-centrifugation studies, the glycoprotein seemed to be predominantly present in the soluble fraction (100000g supernatant). This suggests that it is either largely a soluble cytoplasmic component or is very loosely bound to a membrane, being readily released under the gentlest homogenization procedure. 5. The half-life of Tamm-Horsfall glycoprotein in human kidney was found by the radioimmunoassay method to be approx. 16h. The similarity between the composition of Tamm-Horsfall glycoprotein and human erythropoietin is discussed.

| S-EPMC1166001 | biostudies-other

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