共查询到20条相似文献,搜索用时 31 毫秒
1.
Tamm-Horsfall protein (THP), a normal constituent of mammalian urine, has been determined in rat urine under various conditions in an attempt to elucidate the physiological role of this glycoprotein. Experiments were designed to assess whether THP production is related to the process of urine concentration or to the transport activity of the thick ascending limb of the loop of Henle (TAL), the nephron segment where it is produced. For this purpose, THP excretion was measured, by radioimmunoassay, in adult male rats under 4 different conditions induced by the following chronic treatments: (1) furosemide (12 mg/day in osmotic minipumps); (2) increased water intake; (3) antidiuretic hormone (ADH) infusion (50 ng DDAVP/day in osmotic minipumps) in rats of the Brattleboro strain with hereditary hypothalamic diabetes insipidus; (4) high-protein (32% casein) versus low-protein diet (10% casein). Each experiment included 6 experimental and 6 control rats. After treatment for 1-3 weeks, 24-h urines were collected for determination of urine flow rate, osmolality, and creatinine and THP concentrations. No significant changes in THP excretion were observed in experiments (1) and (2) despite 5- to 7-fold-differences in urine flow rate. Antidiuretic hormone treatment in (3) slightly lowered THP excretion (287 +/- 53 vs. 367 +/- 41 micrograms/day per 100 g body weight; p less than 0.005), whereas high-protein diet, in experiment (4), led to a 50% increase in THP excretion (446 +/- 57 vs. 304 +/- 79 micrograms/day per 100 g body weight; p less than 0.001). Expressing THP excretion relative to that of creatine did not change these findings. These results show (1) that chronically established changes in the level of diuresis, chronic furosemide-induced blockade of the Na,K,Cl-cotransporter or the absence of ADH in Brattleboro rats have little or no impact on the level of THP production, and (2) that THP production is independent of the intensity of transport in the TAL, since two conditions which both are known to increase the transport rate of solutes in the TAL (ADH infusion and high-protein diet), resulted in opposite changes in THP excretion. It is concluded that the rate of THP synthesis is neither linked to the process of urine concentration nor to the ion transport activity of the TAL. 相似文献
2.
As the sugar moiety of Tamm-Horsfall protein (THP) is affected by many pathological conditions, the aim of this study was to examine the influence of carbohydrate-deficient glycoprotein syndrome (CDG) on THP glycans. THP was isolated from urine of one patient with CDG type I and N-glycan profiling, analysis of monosaccharide content, determination of THP reactivity with specific lectins and with anti-THP antibodies were performed. THP of the CDG patient showed markedly lower amounts of all monosaccharides. Diminished amounts of lactosamine-type chains, galactose and alpha2,3 linked sialic acid were expressed in lower reactivity with PHA-L, DSA and MAA, respectively. These modifications were reflected in altered proportions of tetrasialylated and disialylated oligosaccharide chains. THP of the CDG patient reacted slightly more with anti-THP antibodies. Our results indicate that the CDG type I affects the THP sugar moiety and slightly enhances the THP immunoreactivity. 相似文献
3.
Uromodulin (Tamm-Horsfall glycoprotein/uromucoid) is a phosphatidylinositol-linked membrane protein 总被引:10,自引:0,他引:10
M J Rindler S S Naik N Li T C Hoops M N Peraldi 《The Journal of biological chemistry》1990,265(34):20784-20789
Uromodulin, originally identified as an immunosuppressive glycoprotein in the urine of pregnant women, has been previously shown to be identical to human Tamm-Horsfall glycoprotein (THP). THP is synthesized by the kidney and localizes to the renal thick ascending limb and early distal tubule. It is released into the urine in large quantities and thus represents a potential candidate for a protein secreted in a polarized fashion from the apical plasma membrane of epithelial cells in vivo. After introduction of the full-length cDNA encoding uromodulin/THP into HeLa, Caco-2, and Madin-Darby canine kidney cells by transfection, however, the expressed glycoprotein was almost exclusively cell-associated, as determined by immunoprecipitation after radioactive labeling of the cells. By immunofluorescence, THP was localized to the plasma membranes of transfected cells. In transfected cell extracts, THP also remained primarily in the detergent phase in a Triton X-114 partitioning assay, indicating that it has a hydrophobic character, in contrast to its behavior after isolation from human urine. Triton X-114 detergent-associated THP was redistributed to the aqueous phase after treatment of cell extracts with phosphatidylinositol-specific phospholipase C. Treatment of intact transfected HeLa cells with phosphatidylinositol-specific phospholipase C also resulted in the release of THP into the medium, suggesting that it is a glycosylphosphatidylinositol (GPI)-linked membrane protein. Similar to other known GPI-linked proteins, uromodulin/THP contains a stretch of 16 hydrophobic amino acids at its extreme carboxyl terminus which could function as a GPI addition signal and was shown to label with [3H]ethanolamine. The results indicate that THP is a member of this class of lipid-linked membrane proteins and is released into the urine after the loss of its hydrophobic anchor, probably by the action of a phospholipase or protease. 相似文献
4.
Tamm-Horsfall protein (THP), also known as uromodulin, is a major glycoprotein synthesized in the kidney. THP is expressed on the luminal surface of the membrane with the glycosyl phosphatidylinositol (GPI) anchor and excreted in urine at a rate of 50-100 mg per day. Although THP is the most abundant urinary protein, the function of THP remains unclear. In addition, little is known about the mechanism by which large amounts of THP are actively released into the urinary fluid. In this study, we examined the C-terminal structure of highly purified THP derived from human urine. Carboxypeptidase Y efficiently degraded urinary THP, indicating that the C-terminal structure of the protein contains an amino acid residue with a free carboxyl moiety. These results are consistent with our previous finding that urinary THP does not bind anti-CRD antibody. We obtained peptides from the complete digestion of urinary THP with lysylendopeptidase. We purified the most C-terminal peptide with p-phenylene diisothiocyanate-controlled pore glass (DITC-CPG) beads. N-terminal sequence analysis indicated the peptide begins with Tyr 520 and ends between E539 and E576. Direct C-terminal amino acid sequencing of highly purified urinary THP gave a sequence of -X-(Q)-G-(R)-F, corresponding to amino acids 544-548, -S-Q-G-R-F. We therefore conclude that urinary THP is generated by a proteolytic cleavage between F548 and S549, 66 amino acids upstream of a possible GPI-anchor attachment site. Because the sequence of THP, including the cleavage site, is highly homologous to that of GP2, a GPI-anchored protein within the pancreas, and both THP and GP2 are abundantly found as soluble forms in the excreted fluids, a common mechanism may exist governing the proteolytic release of GPI-anchored membrane proteins. 相似文献
5.
Zhanying Hong Jun Wen Quanlong Zhang Guorong Fan Yifeng Chai Yutian Wu 《Chirality》2010,22(3):355-360
The objective of this work was to study the stereoselectivity in excretion of tetrahydropalmatine (THP) enantiomers by rats and identify the metabolites of racemic THP (rac‐THP) in rat urine. Urine and bile samples were collected at various time intervals after a single oral dose of rac‐THP. The concentrations of THP enantiomers in rat urine and bile were determined using a modification of an achiral–chiral high‐performance liquid chromatographic (HPLC) method that had been previously published. The cumulative urinary excretion over 96 h of (?)‐THP and (+)‐THP was found to be 55.49 ± 36.9 μg and 18.33 ± 9.7 μg, respectively. The cumulative biliary excretion over 24 h of (?)‐THP and (+)‐THP was 19.19 ± 14.6 μg and 12.53 ± 10.4 μg, respectively. The enantiomeric (?/+) concentration ratios of THP changed from 2.80 to 5.15 in urine, and from 1.36 to 1.80 in bile. The mean cumulative amount of (?)‐THP was significantly higher than that of (+)‐THP both in urine and bile samples. However, the enantiomeric (?/+) concentration ratios in rat urine and bile were significantly lower than those ratios in rat plasma. These findings suggested the excretion of THP enantiomers was stereoselective rather than a reflection of chiral pharmacokinetic aspects in plasma and (?)‐THP was preferentially excreted in rat urine and bile. Three O‐demethylation metabolites and the parent drug rac‐THP were detected by liquid chromatography‐tandem mass spectrometry in rat urine. One metabolite was obtained by preparative HPLC and identified as 10‐O‐demethyl‐THP. Chirality, 2010. © 2009 Wiley‐Liss, Inc. 相似文献
6.
Rabbit urinary tamm-horsfall glycoprotein. Chemical composition and tentative carbohydrate structure
The urinary Tamm-Horsfall protein (THP) is the major glycoprotein secreted by the mammalian kidney. We recently isolated and immortalized thick ascending limb of Henle cells from rabbit kidney, which produce Tamm-Horsfall protein in cell culture in vitro. In order to further study the yet undefined functional role and biosynthetic pathways of this protein, we first re-examined the chemical composition and the carbohydrate structure of rabbit urinary Tamm-Horsfall protein. Using precipitation with 0.58 mol/l NaCl a protein was isolated from rabbit urine which showed extensive microheterogeneity and had an average molecular mass of 95 kDa. Deglycosylation of the protein led to a loss of microheterogeneity and yielded a molecular mass of 58-60 kDa. Amino-acid analysis of the native and deglycosylated protein revealed a lower cysteine (20 mol/mol THP) and a higher histidine (20 mol/mol THP) content than described previously. Chemical analysis of the carbohydrates showed a high glucosamine (50 mol/mol THP), galactose (43 mol/mol THP), and mannose (24 mol/mol THP) content. The amount of sialic acid was 15 mol/mol THP. Using lectins to identify the structure of the carbohydrate chains it was shown that rabbit Tamm-Horsfall protein possesses complex-type oligosaccharide chains with terminal sialic acid, beta-galactose, and probably alpha-fucose and chains of the mucin type. These results indicate that some of the cysteine residues in the polypeptide chain of THP can be replaced by histidine, suggesting a role of some cysteins in metal binding rather than intramolecular stabilization.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
7.
Cavallone D Malagolini N Serafini-Cessi F 《Biochemical and biophysical research communications》2001,280(1):110-114
Human Tamm-Horsfall glycoprotein (THP) is synthesised in the thick ascending limb of Henle and convoluted distal tubules, inserted into luminal cell-surface by the glycosyl-phosphatidylinositol (GPI)-anchor and excreted in urine at a rate of 50-100 mg per day. Up to date there is no indication on the way in which THP is excreted into the urinary fluid. In this study, we examined by Western blotting THP from human kidney in comparison to urinary THP. As expected for a GPI-anchored protein, THP was recovered from the kidney lysate in a Triton X-100 insoluble form, which moved in a sucrose gradient to a zone of low density. The apparent molecular weight of kidney THP appeared greater than that of urinary THP, but no difference in the electrophoretic mobility was observed when the former was subjected to GPI-specific phospholipase-C treatment, strongly suggesting that a proteolytic cleavage at the juxtamembrane-ectodomain of kidney THP is responsible for the urinary excretion. 相似文献
8.
9.
Rhodes DC 《Immunology and cell biology》2000,78(5):474-482
The purpose of the present study was to quantify the binding affinity between Tamm-Horsfall protein (THP) and complement 1q (C1q) using ELISA and a resonant mirror biosensor. In ELISA, immobilized THP was incubated with soluble C1q under both low and physiological ionic-strength conditions. Tamm-Horsfall protein bound C1q with an equilibrium dissociation constant (KD) of 1.9 +/- 0.6 nmol/L in low ionic-strength Tris buffers (20 mmol/L NaCl, pH 7.5) and with a lower affinity (KD of 13.4 +/- 4.7 nmol/L) in physiological-strength Tris buffers (154 mmol/L NaCl, pH 7.5). A resonant mirror biosensor, which monitors binding events in real-time, was used to quantify the KD of this reaction, as well as to estimate the kinetic parameters. In these studies, THP and C1q bound with an association rate constant, kass, of 1.25 x 105 L/mol per s and a dissociation rate constant, kdiss, of 0.002-0.005/s. The calculated KD for the THP/C1q binding in low ionic-strength buffers was higher (averages of 10-15 nmol/L) than that obtained by the ELISA, while physiological ionic-strength buffers still reduced the affinity of this binding by an order of magnitude. In conclusion, THP consistently bound C1q with high affinity using several techniques. At least a portion of this interaction involved electrostatic events, as demonstrated by the influence of ionic strength on the binding affinity. 相似文献
10.
Tissue-specific expression of human genes and secretion of human proteins into the body fluids in transgenic animals provides an important means of manufacturing large-quantity and high-quality pharmaceuticals. The present study demonstrates using transgenic mice that a 3.0 kb promoter of the mouse Tamm-Horsfall protein (THP, or uromodulin) gene directs the specific expression of human growth hormone (hGH) gene in the kidney followed by the secretion of hGH protein into the urine. hGH expression was detected in renal tubules that actively produce the THP, that is, the ascending limb of Henle's loop and distal convoluted tubules. Up to 500 ng/ml of hGH was detected in the urine, and this level remained constant throughout the 10-month observation period. hGH was also detectable in the stomach epithelium and serum in two of the transgenic lines, suggesting position-dependent effects of the transgene and leakage of hGH from the site of synthesis into the bloodstream, respectively. These results indicate that the 3.0 kb mouse THP promoter is primarily kidney-specific and can be used to convert kidney into a bioreactor in transgenic animals to produce recombinant proteins. Given the capacity of urine production independent of age, sex and lactation, the ease of urinary protein purification, and the potentially distinct machinery for post-translational modifications in the kidney epithelial cells, the kidney-based transgenic bioreactor may offer unique opportunities for producing certain complex pharmaceuticals. 相似文献
11.
Synthesis and processing of thionin precursors in developing endosperm from barley (Hordeum vulgare L.) 总被引:2,自引:1,他引:1
下载免费PDF全文
![点击此处可从《The EMBO journal》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Ponz F Paz-Ares J Hernández-Lucas C Carbonero P García-Olmedo F 《The EMBO journal》1983,2(7):1035-1040
Thionin is a lysine-rich polypeptide (mol. wt. 5000) which is synthesized in developing barley endosperm from ˜8 days to ˜30 days after anthesis. Two thionin precursors (THP1 and THP2) have been identified using monospecific antibodies (A-TH) prepared against the mature protein. THP1, which is the only polypeptide recognized in vitro by A-TH, is encoded by a 7.5S mRNA obtained from membrane-bound polysomes, and its alkylated derivative has an apparent mol. wt. of 17 800. THP2, which is selected together with mature thionin by A-TH among labelled proteins in vivo, differs from THP1 in apparent mol. wt. (17 400 alkylated) and in electrophoretic mobility at pH 3.2. Both THP1 and THP2 are competed out of the antigen-antibody complex by purified thionin. The conversion of THP2 into thionin, which has been demonstrated in a pulse-chase experiment in vivo, is a post-translational process. As it has not been possible to detect THP1 in vivo it is assumed that it is converted co-translationally into THP2. Final deposition of thionin as an extrinsic membrane protein, possibly associated with the endoplasmic reticulum, has been tentatively established on the basis of subcellular fractionation experiments. 相似文献
12.
Alexander J. Howie Christopher J. Lote Andrew N. Hillis 《Cell and tissue research》1991,263(3):585-587
Summary Tamm-Horsfall protein (THP) is the main protein in normal human urine, and is found in the thick limb of the Loop of Henle in human kidney, and in other mammalian species. The skin of the frog, Rana temporaria, has similar physiological properties to this mammalian kidney tissue. In the present study, an immunohistological method involving an antibody to human THP was used to investigate the distribution of this distinctive protein in frog kidney and skin, and to compare its distribution with that found in the kidney tubules of rat and rabbit. THP-positive material was detected in the distal renal tubules and nephric duct of frogs, and was also located in the superificial epidermis of skin. It is suggested that its presence in amphibian skin is consistent with the hypothesis that THP is an important component of tissues that absorb sodium and chloride ions, but remain impermeable to water. 相似文献
13.
V T Andriole 《The Yale journal of biology and medicine》1985,58(2):91-100
Recurrent bacterial infection of the kidney was previously thought to be responsible for the renal scarring typical of chronic pyelonephritis until recent studies suggested that recurrent bacteriuria rarely produces chronic pyelonephritis in the absence of obstructive uropathy. In contrast, the association between vesicoureteral reflux (VUR) and chronic pyelonephritis has been observed frequently in the absence of urinary infection. Although the mechanism by which VUR injures the kidney has not been defined, recent observations have suggested that some component of urine might serve as an antigenic determinant involved in the immunopathogenesis of renal scarring in VUR. Therefore, the present studies investigated the immunopathogenic role of Tamm-Horsfall protein (THP) in (1) a rabbit model of tubulointerstitial nephritis; (2) a swine model of reflux nephropathy; and (3) patients with recurrent nephrolithiasis. The antigenic similarities between THP and uropathic bacteria were also studied. Our observations indicate that autoimmune responses to THP may occur after exposure to THP by intravenous challenge in rabbits, by urinary reflux in pigs, and in recurrent nephrolithiasis in man. Also, extracts of uropathic coliforms competitively inhibit the binding of human THP to its antibody. These studies suggest that autoimmune responses to THP may be the pathogenetic mechanism by which these factors, including bacteriuria, contribute to "chronic pyelonephritis." 相似文献
14.
In an attempt to isolate a heparanase receptor, postulated to mediate non-enzymatic functions of the heparanase protein, we utilized human urine collected from healthy volunteers. Affinity chromatography of this rich protein source on immobilized heparanase revealed resistin as a heparanase binding protein. Co-immunoprecipitation and ELISA further confirmed the interaction between heparanase and resistin. Importantly, we found that heparanase potentiates the bioactivity of resistin in its standard bioassay in which monocytic human leukemia cell line, THP1, differentiates into adherent macrophage-like foam cells. It is thus conceivable that this newly identified complex of heparanase and resistin exerts a stimulatory effect also in various inflammatory conditions known to be affected by the two proteins. 相似文献
15.
Tamm-Horsfall glycoprotein (THP) is the most abundant protein which is synthesized by renal tubular cells and excreted in urine. Its role in urinary tract infection has yet not been identified. In the present study, the contribution of THP towards adherence of Pseudomonas aeruginosa to uroepithelial cells and murine peritoneal macrophages was studied. Decreased adherence of THP-coated P. aeruginosa to UECs and phagocytes was observed in vitro. In vivo, P. aeruginosa showed increased renal bacterial load and tissue pathology in a mouse model of acute ascending pyelonephritis, when THP-coated P. aeruginosa was used to cause infection. This study shows that THP may not necessarily act as a host defense component; rather, it may help in renal colonization of P. aeruginosa in vivo. 相似文献
16.
Rahul Mittal Saroj Sharma Sanjay Chhibber Kusum Harjai 《World journal of microbiology & biotechnology》2006,22(9):915-919
Summary Tamm-Horsfall glycoprotein is the most abundant protein in human urine. The present investigation was planned to study the effect of Tamm-Horsfall protein (THP) on elaboration of virulence factors by biofilm cells of Pseudomonas aeruginosa. It was observed that with increase in concentration of THP from 10 to 50 μg/ml there was significant enhancement in elaboration of all the virulence factors by biofilm cells of P. aeruginosa. However, with further increase in concentration of THP from 50 to 70 μg/ml, significant decrease in elaboration of all the virulence traits was observed. Implications of these findings in relation to urinary tract infections caused by P. aeruginosa have been discussed. 相似文献
17.
Shen HL Xu ZG Huang LY Liu D Lin DH Cao JB Zhang X Wang ZQ Wang WH Yang PY Han ZG 《Molecular and cellular biochemistry》2009,321(1-2):73-83
Liver-specific ZP domain-containing protein (LZP) was recently identified as a secreted protein that is specifically expressed in liver. However, the physiological effects of LZP are largely unknown. In this study, we found that LZP was detectable in mouse kidneys, testes, ovaries and heart, in addition to liver. LZP was localized in the spermatid cells of testes, corpus luteum cells of ovaries, and cardiac muscle cells of heart. But the protein mainly anchored on the apical membrane of the thick ascending limb of the loop of Henle (TAL) cell in mouse kidney. In rat kidney LZP and Tamm-Horsfall protein (THP) were co-localized in TAL. The in vivo interaction between LZP and THP was confirmed in kidney and urine by co-immunoprecipitation assay, and the in vitro interaction was detected by GST pull-down assay, implying that the interaction could be independent on N-linked glycosylated modification of LZP. Surprisingly, LZPs with intramolecular disulfide bridges could self-interact, and then self-aggregate into spheres of varying sizes, but not polymerize into filaments. The finding that LZP might act as a new partner of THP would provide novel insights into renal functions related to THP and LZP, such as the urothelial permeability barrier and the host defense against the adhesion of pathogens. 相似文献
18.
Easton RL Patankar MS Clark GF Morris HR Dell A 《The Journal of biological chemistry》2000,275(29):21928-21938
Tamm-Horsfall glycoprotein (THP) is a major glycoprotein associated with human urine that binds pro-inflammatory cytokines and also inhibits in vitro T cell proliferation induced by specific antigens. THP derived from human pregnancy urine (designated uromodulin) has previously been shown to be 13-fold more effective as an inhibitor of antigen-induced T cell proliferation than THP obtained from other sources. Structural analysis of human THP and uromodulin has for the first time revealed that these glycoproteins are O-glycosylated. THP from nonpregnant females and males expresses primarily core 1 type O-glycans terminated with either sialic acid or fucose but not the sialyl Lewis(x) epitope. By contrast, the O-glycans linked to uromodulin include unusual core 2 type glycans terminated with one, two, or three sialyl Lewis(x) sequences. The specific association of these unusual carbohydrate sequences with uromodulin could explain its enhanced immunomodulatory effects compared with THP obtained from males and nonpregnant females. Analysis of THP from one of the pregnant females 2 months postpartum showed a reversion of the O-glycan profile to that found for a non-pregnant female. These data suggest that the glycosylation state of uromodulin could be under the regulation of steroidal hormones produced during pregnancy. The significant physiological implications of these observations are discussed. 相似文献
19.
The chloroethoxyethyl (CEE) group is completely stable under the acidic conditions required to remove the 5'-protecting groups in the oligoribonucleotide synthesis, but can be cleaved under the similar condition to that of the tetrahydropyranyl (THP) group in region of pH 2-3. The oligoribonucleotides were synthesized by the phosphoramidite method on solid supports. 相似文献
20.
Jesse L. Cashaw 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》1993,613(2)
Tetrahydropapaveroline (THP) concentrations were measured in the urine of Parkinsonian patients receiving l-dopa—carbidopa (Sinemet) therapy, using a method that employs a separation scheme that selectively isolates THP from urine and utilizes the Pictet—Spengler condensation of THP with formaldehyde combined with high-performance liquid chromatography for identification and determination. The mean (± S.D.) recoveries of THP from normal urine with 0.2 pmol/ml added and from Parkinsonian patients' urines with 0.5 pmol/ml added were 48.6 ± 5.7 and 44.6 ± 3.1%, respectively. Three Parkinsonian patients who were receiving either 250, 750 or 1000 mg of l-dopa (as Sinemet) daily had 24-h urinary THP excretion levels of 989, 1017 and 1600 pmol, respectively. 相似文献