scholarly journals Some physical and immunological properties of ox kidney biliverdin reductase

1988 ◽  
Vol 255 (2) ◽  
pp. 431-435 ◽  
Author(s):  
E M Rigney ◽  
O Phillips ◽  
T J Mantle
Keyword(s):  
Rat Kidney ◽  
Proximal Tubules ◽  
Monomeric Form ◽  
Thiol Groups ◽  
Glutathione S Transferase ◽  
Free Concentration ◽  
Biliverdin Reductase ◽  
Kidney Enzyme ◽  
Intracellular Binding ◽  
Inner Cortex

The liver, kidney and spleen of the mouse and rat and the kidney and spleen of the ox express a monomeric form of biliverdin reductase (Mr 34,000), which in the case of the ox kidney enzyme exists in two forms (pI 5.4 and 5.2) that are probably charge isomers. The livers of the mouse and rats express, in addition, a protein (Mr 46,000) that cross-reacts with antibodies raised against the ox kidney enzyme and may be related to form 2 described by Frydman, Tomaro, Awruch & Frydman [(1983) Biochim. Biophys. Acta 759, 257-263]. Higher-Mr forms appear to exist in the guinea pig and hamster. The ox kidney enzyme has three thiol groups, of which two are accessible to 5,5′-dithiobis-(2-nitrobenzoate) in the native enzyme. Immunocytochemical analysis reveals that biliverdin reductase is localized in proximal tubules of the inner cortex of the rat kidney. Biliverdin reductase antiserum also stains proximal tubules in human and ox kidney. The staining of podocytes in glomeruli of ox kidney with antiserum to aldose reductase is particularly prominent. The localization of biliverdin reductase in the inner cortical zone of rat kidney is similar to that described for glutathione S-transferase YfYf, and it is suggested that one function of this ‘intracellular binding protein’ may be to maintain a low free concentration of biliverdin to allow biliverdin reductase to operate efficiently.

scholarly journals Cloning and overexpression of rat kidney biliverdin IXα reductase as a fusion protein with glutathione S-transferase: stereochemistry of NADH oxidation and evidence that the presence of the glutathione S-transferase domain does not effect BVR-A activity

1997 ◽  
Vol 328 (1) ◽  
pp. 33-36 ◽  
Author(s):  
Orla ENNIS ◽  
Robin MAYTUM ◽  
J. Timothy MANTLE
Keyword(s):  
Fusion Protein ◽  
Initial Rate ◽  
Rat Kidney ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Glutathione S Transferase ◽  
Catalysed Oxidation ◽  
Kidney Enzyme ◽  
Presence And Absence ◽  
Burst Kinetics

Native biliverdin IXα reductase (BVR-A) is a monomer of molecular mass 34 kDa. We have developed an expression vector that allows the isolation of 40 mg of a glutathione S-transferase (GST)-BVR-A fusion protein from 1 litre of culture. The fusion protein (60 kDa) behaves as a dimer on gel filtration (120 kDa), so that we have artificially created a BVR-A dimer. The recombinant rat kidney enzyme exhibits pre-steady-state ‘burst’ kinetics that show a pH dependence similar to that already described for ox kidney BVR-A. Similar behaviour was obtained in the presence and absence of the GST domain both for the burst kinetics and during initial-rate studies in the presence and absence of albumin. The stereospecificity of the BVR-A-catalysed oxidation of [4-3H]NADH, labelled at the A and B faces, was shown to occur exclusively via the B face.

scholarly journals ON THE LOCALIZATION OF SOME PHOSPHATASES IN THREE DIFFERENT SEGMENTS OF THE PROXIMAL TUBULES IN THE RAT KIDNEY

1967 ◽  
Vol 15 (8) ◽  
pp. 456-469 ◽  
Author(s):  
N. O. JACOBSEN ◽  
F. JØRGENSEN ◽  
Å. C. THOMSEN
Keyword(s):  
Adenosine Triphosphate ◽  
Rat Kidney ◽  
Adenosine Monophosphate ◽  
Adenosine Diphosphate ◽  
Proximal Tubules ◽  
Reaction Pattern ◽  
Cytoplasmic Bodies ◽  
Acid And Alkaline Phosphatase ◽  
Convoluted Tubules

The distribution of several phosphatases in three segments of the proximal tubules was studied in frozen sections of glutaraldehyde-fixed rat kidneys. Two segments of the convoluted tubules were identified by in vivo injection of trypan blue. By increasing the concentration of adenosine triphosphate to 3 mM in the Wachstein and Meisel ATPase medium, a clear segmental differentiation in the reaction pattern of the brush border, cytoplasmic bodies and basal infoldings of the proximal tubules was obtained. The specificity of the reaction was investigated by substituting adenosine diphosphate, adenosine monophosphate or β-glycerophosphate for adenosine triphosphate in the incubation medium and by employing cyanide or fluoride as inhibitors. The reaction pattern was also compared with the localization of acid and alkaline phosphatase activities. In addition, the distribution of glucose 6-phosphatase activity was studied which showed differences in the three segments of the proximal tubules.

scholarly journals Purification and properties of 2-aminoadipate: 2-oxoglutarate aminotransferase from bovine kidney

1989 ◽  
Vol 261 (3) ◽  
pp. 761-768 ◽  
Author(s):  
D R Deshmukh ◽  
S M Mungre
Keyword(s):  
Rat Kidney ◽  
Deae Cellulose ◽  
Dicarboxylic Acids ◽  
Structural Similarity ◽  
Appreciable Amount ◽  
Kinetic Properties ◽  
Bovine Kidney ◽  
Purification And Properties ◽  
Kidney Enzyme ◽  
Structural Differences

Previous studies with rat kidney preparations indicated that 2-aminoadipate aminotransferase (AadAT) and kynurenine aminotransferase (KAT) activities are properties of a single protein. We found that bovine kidney contains an appreciable amount of AadAT activity, but lacks KAT activity. AadAT from bovine and rat kidney extracts were purified to electrophoretic homogeneity. The purification procedure included fractionation with (NH1)2SO1, heat treatment, DEAE-cellulose chromatography and hydroxyapatite chromatography. Physical and kinetic properties, such as pH optima, Km for substrates, Mr, electrophoretic mobility and inhibition by dicarboxylic acids of bovine kidney AadAT, were similar to those of the rat kidney enzyme. However, bovine kidney AadAT differed from rat kidney AadAT in substrate specificity, amino acid composition and stability when stored. The titration curve of bovine kidney AadAT was also different from that of the rat kidney enzyme. The results suggest that bovine kidney AadAT may have some structural similarity to rat kidney AadAT and that the structural differences observed between the two enzymes may explain the absence of KAT activity in bovine kidney.

Sepsis induces changes in the expression and distribution of Toll-like receptor 4 in the rat kidney

2006 ◽  
Vol 290 (5) ◽  
pp. F1034-F1043 ◽  
Author(s):  
Tarek M. El-Achkar ◽  
Xiaoping Huang ◽  
Zoya Plotkin ◽  
Ruben M. Sandoval ◽  
Georges J. Rhodes ◽  
...  
Keyword(s):  
Rat Kidney ◽  
Expression Patterns ◽  
Cecal Ligation ◽  
Proximal Tubules ◽  
Microbial Pathogens ◽  
Sprague Dawley ◽  
Renal Vasculature ◽  
Two Photon Microscopy ◽  
Sprague Dawley Rats ◽  
Cellular Signal

Toll-like receptors (TLRs) are now recognized as the major receptors for microbial pathogens on cells of the innate immune system. Recently, TLRs were also identified in many organs including the kidney. However, the cellular distribution and role of these renal TLRs remain largely unknown. In this paper, we investigated the expression of TLR4 in a cecal ligation and puncture (CLP) model of sepsis in Sprague-Dawley rats utilizing fluorescence microscopy. In sham animals, TLR4 was expressed predominantly in Tamm-Horsfall protein (THP)-positive tubules. In CLP animals, TLR4 expression increased markedly in all tubules (proximal and distal), glomeruli, and the renal vasculature. The staining showed a strong apical distribution in all tubules. A moderately less intense cellular signal colocalized partially with the Golgi apparatus. In addition, kidneys from septic rats showed increased expression of CD14 and THP. They each colocalized strongly with TLR4, albeit in different tubular segments. We also imaged the kidneys of live septic animals with two-photon microscopy after fluorescent lipopolysaccharide (LPS) injection. Within 10 min, LPS was seen at the brush border of some proximal tubules. Within 60 min, LPS was fully cytoplasmic in proximal tubules. Conversely, distal tubules showed no LPS uptake. We conclude that TLR4, CD14, and THP have specific renal cellular and tubular expression patterns that are markedly affected by sepsis. Systemic endotoxin can freely access the tubular and cellular sites where these proteins are present. Therefore, locally expressed TLRs and other interacting proteins could potentially modulate the renal response to systemic sepsis.

Identification of an apical \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{Cl}^{-}{/}\mathrm{HCO}_{3}^{-}\) \end{document} exchanger in rat kidney proximal tubule

2003 ◽  
Vol 285 (3) ◽  
pp. C608-C617 ◽  
Author(s):  
Snezana Petrovic ◽  
Liyun Ma ◽  
Zhaohui Wang ◽  
Manoocher Soleimani
Keyword(s):  
Proximal Tubule ◽  
Apical Membrane ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Immunocytochemical Staining ◽  
Kidney Cortex ◽  
Rat Kidney Cortex ◽  
Kidney Proximal Tubule ◽  
Anion Transporter

SLC26A6 (or putative anion transporter 1, PAT1) is located on the apical membrane of mouse kidney proximal tubule and mediates [Formula: see text] exchange in in vitro expression systems. We hypothesized that PAT1 along with a [Formula: see text] exchange is present in apical membranes of rat kidney proximal tubules. Northern hybridizations indicated the exclusive expression of SLC26A6 (PAT1 or CFEX) in rat kidney cortex, and immunocytochemical staining localized SLC26A6 on the apical membrane of proximal tubules, with complete prevention of the labeling with the preadsorbed serum. To examine the functional presence of apical [Formula: see text] exchanger, proximal tubules were isolated, microperfused, loaded with the pH-sensitive dye BCPCF-AM, and examined by digital ratiometric imaging. The pH of the perfusate and bath was kept at 7.4. Buffering capacity was measured, and transport rates were calculated as equivalent base flux. The results showed that in the presence of basolateral DIDS (to inhibit [Formula: see text] cotransporter 1) and apical EIPA (to inhibit Na+/H+ exchanger 3), the magnitude of cell acidification in response to addition of luminal Cl– was ∼5.0-fold higher in the presence than in the absence of [Formula: see text]. The Cl–-dependent base transport was inhibited by ∼61% in the presence of 0.5 mM luminal DIDS. The presence of physiological concentrations of oxalate in the lumen (200 μM) did not affect the [Formula: see text] exchange activity. These results are consistent with the presence of SLC26A6 (PAT1) and [Formula: see text] exchanger activity in the apical membrane of rat kidney proximal tubule. We propose that SLC26A6 is likely responsible for the apical [Formula: see text] (and Cl–/OH–) exchanger activities in kidney proximal tubule.

Immunolocalization of NHE8 in rat kidney

2005 ◽  
Vol 288 (3) ◽  
pp. F530-F538 ◽  
Author(s):  
Sunita Goyal ◽  
SueAnn Mentone ◽  
Peter S. Aronson
Keyword(s):  
Proximal Tubule ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Rat Kidney ◽  
Surface Membrane ◽  
Proximal Tubules ◽  
Intense Staining ◽  
Proximal Tubule Cells ◽  
Nephron Segment ◽  
Tubule Cells

In situ hybridization studies demonstrated that Na+/H+ exchanger NHE8 is expressed in kidney proximal tubules. Although membrane fractionation studies suggested apical brush-border localization, precise membrane localization could not be definitively established. The goal of the present study was to develop isoform-specific NHE8 antibodies as a tool to directly establish the localization of NHE8 protein in the kidney by immunocytochemistry. Toward this goal, two sets of antibodies that label different NHE8 epitopes were developed. Monoclonal antibody 7A11 and polyclonal antibody Rab65 both specifically labeled NHE8 by Western blotting as well as by immunofluorescence microscopy. The immunolocalization pattern in the kidney seen with both antibodies was the same, thereby validating NHE8 specificity. In particular, NHE8 expression was observed on the apical brush-border membrane of all proximal tubules from S1 to S3. The most intense staining was evident in proximal tubules in the deeper cortex and medulla with a significant but somewhat weaker staining in superficial proximal tubules. Colocalization studies with γ-glutamyltranspeptidase and megalin indicated expression of NHE8 on both the microvillar surface membrane and the coated-pit region of proximal tubule cells, suggesting that NHE8 may be subject to endocytic retrieval and recycling. Although colocalizing in the proximal tubule with NHE3, no significant alteration in NHE8 protein expression was evident in NHE3-null mice. We conclude that NHE8 is expressed on the apical brush-border membrane of proximal tubule cells, where it may play a role in mediating or regulating ion transport in this nephron segment.

Ouabain-insensitive, Na-ATPase activity in pure suspensions of rat kidney proximal tubules

FEBS Letters ◽  
1990 ◽  
Vol 269 (1) ◽  
pp. 77-78 ◽  
Author(s):  
Reinaldo Marín ◽  
Daniela C. Gómez ◽  
Gloria A. Rodríguez ◽  
Teresa Proverbio ◽  
Fulgencio Proverbio
Keyword(s):  
Atpase Activity ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Kidney Proximal Tubules

scholarly journals Immunoelectron Microscopic Localization of the Electrogenic Na/HCO3 Cotransporter in Rat and Ambystoma Kidney

2000 ◽  
Vol 11 (12) ◽  
pp. 2179-2189
Author(s):  
ARVID B. MAUNSBACH ◽  
HENRIK VORUM ◽  
TAE-HWAN KWON ◽  
SØREN NIELSEN ◽  
BRIAN SIMONSEN ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Proximal Tubule ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Kidney Cortex ◽  
Apical Plasma Membrane ◽  
Rat Kidney Cortex ◽  
C Terminus ◽  
Basolateral Plasma Membrane ◽  
Intracellular Vesicles

Abstract. Immunofluorescence analysis has revealed that electrogenic Na+/HCO3- (NBC1) is expressed in the proximal tubule of rat kidney and in the proximal and distal tubules of the salamander Ambystoma tigrinum kidney. The present study was undertaken to define the detailed subcellular localization of the NBC1 in rat and Ambystoma kidney using high-resolution immunoelectron microscopy. For this purpose, two rabbit polyclonal antibodies raised against amino acids 928 to 1035 and amino acids 1021 to 1035 of the C-terminus of rat kidney (rkNBC1) were developed. The affinity-purified antibodies revealed a strong band of approximately 140 kD in immunoblots of membranes from rat kidney cortex but no signal in membranes isolated from outer and inner medulla. Deglycosylation reduced the apparent molecular weight to approximately 120 kD, corresponding to the predicted molecular weight. A similar but weaker band was also present in membranes isolated from the lateral part of Ambystoma kidney. In rat kidney, immunohistochemistry confirmed the presence of rkNBC1 in convoluted segments of the proximal tubules. In ultrathin cryosections or Lowicryl HM20 sections from rat kidney cortex, distinct immunogold labeling was associated with the basolateral plasma membrane of segments S1 and S2 of proximal tubules, whereas in S3 no labeling was observed. The labeling density was similar at the basal and lateral plasma membrane and was specifically associated with the inner surface of the membrane consistent with the internal position of the C-terminus of the transporter. In contrast, rkNBC1 was absent from the apical plasma membrane and not observed in intracellular vesicles, including those closely associated with basolateral plasma membrane. In Ambystoma kidney, a weak labeling was present in the basolateral membrane of the proximal tubule and stronger labeling was observed in the late distal segment. The results demonstrate that rkNBC1 is expressed only in segment S1 and segment S2 of rat proximal tubule as well as Ambystoma proximal and late distal tubule and that rkNBC1 is present in both basal and lateral plasma membranes and absent in intracellular vesicles of the apical plasma membrane.

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