Characterization of mouse urea transporters UT-A1 and UT-A2

2002 ◽  
Vol 283 (4) ◽  
pp. F817-F825 ◽  
Author(s):  
R. A. Fenton ◽  
G. S. Stewart ◽  
B. Carpenter ◽  
A. Howorth ◽  
E. A. Potter ◽  
...  
Keyword(s):  
Northern Blot Analysis ◽  
Mouse Kidney ◽  
Xenopus Laevis Oocytes ◽  
Loop Of Henle ◽  
Kidney Medulla ◽  
Collecting Ducts ◽  
Terminal Amino ◽  
Type 3

Specialized transporter proteins that are the products of two closely related genes, UT-A ( Slc14a2) and UT-B ( Slc14a1), modulate the movement of urea across cell membranes. The purpose of this study was to characterize the mouse variants of two major products of the UT-A gene, UT-A1 and UT-A2. Screening a mouse kidney inner medulla cDNA library yielded 4,047- and 2,876-bp cDNAs, the mouse homologues of UT-A1 and UT-A2. Northern blot analysis showed high levels of UT-A mRNAs in kidney medulla. UT-A transcripts were also present in testes, heart, brain, and liver. Immunoblots with an antiserum raised to the 19 COOH-terminal amino acids of rat UT-A1 (L194) identified immunoreactive proteins in kidney, testes, heart, brain, and liver and showed a complex pattern of differential expression. Relative to other tissues, kidney and brain had the highest levels of UT-A protein expression. In kidney sections, immunostaining with L194 revealed immunoreactive proteins in type 1 (short) and type 3 (long) thin descending limbs of the loop of Henle and in the middle and terminal inner medullary collecting ducts. Expression in Xenopus laevis oocytes showed that, characteristic of UT-A family members, the cDNAs encoded phloretin-inhibitable urea transporters. Acute application of PKA agonists (cAMP/forskolin/IBMX) caused a significant increase in UT-A1- and UT-A3-, but not UT-A2-mediated, urea transport.

scholarly journals Characterization of cDNAs encoding isoforms of hamster 3 beta-hydroxysteroid dehydrogenase/delta 5-->4 isomerase

1998 ◽  
Vol 20 (1) ◽  
pp. 99-110 ◽  
Author(s):  
FM Rogerson ◽  
J Courtemanche ◽  
A Fleury ◽  
JG LeHoux ◽  
JI Mason ◽  
...  
Keyword(s):  
Hydroxysteroid Dehydrogenase ◽  
Cdna Libraries ◽  
Sexually Dimorphic ◽  
High Affinity ◽  
Liver And Kidney ◽  
Low Levels ◽  
Type 3

Western blot analyses of various hamster tissues reveal high levels of expression of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) in adrenal and liver, and moderate levels of expression in kidney. The expression in liver is sexually dimorphic; very high levels of protein are observed in adult male liver but very low levels are seen in the female liver. Three distinct cDNAs encoding isoforms of 3 beta-HSD were isolated from hamster cDNA libraries. The type 1 isoform is a high-affinity dehydrogenase/isomerase expressed in adrenal and male kidney. The type 2 isoform is also a high-affinity dehydrogenase/isomerase expressed in kidney and male liver. The type 3 enzyme is a 3-ketosteroid reductase expressed predominantly in kidney. Sequencing of the clones showed that all three are structurally very similar, although types 1 and 2 share the greatest degree of similarity. Immunohistochemical staining for 3 beta-HSD in the adrenal was found throughout the adrenal cortex. In the kidney staining was confined to tubules, and in the liver, heavy staining was found in hepatocytes. The cloning of cDNAs for 3 beta-HSD from the liver and kidney should help in elucidating the function of this enzyme in these tissues.

scholarly journals Novel LinA Type 3 δ-Hexachlorocyclohexane Dehydrochlorinase

2015 ◽  
Vol 81 (21) ◽  
pp. 7553-7559 ◽  
Author(s):  
Nidhi Shrivastava ◽  
Zbynek Prokop ◽  
Ashwani Kumar
Keyword(s):  
Amino Acid ◽  
Primary Structure ◽  
Degradation Pathway ◽  
Amino Acid Residues ◽  
Hch Isomers ◽  
New Variant ◽  
Type 3

ABSTRACTLinA is the first enzyme of the microbial degradation pathway of a chlorinated insecticide, hexachlorocyclohexane (HCH), and mediates the dehydrochlorination of α-, γ-, and δ-HCH. Its two variants, LinA type 1 and LinA type 2, which differ at 10 out of 156 amino acid residues, have been described. Their activities for the metabolism of different HCH isomers differ considerably but overall are high for γ-HCH, moderate for α-HCH, low for δ-HCH, and lacking for β-HCH. Here, we describe the characterization of a new variant of this enzyme, LinA type 3, whose gene was identified from the metagenome of an HCH-contaminated soil sample. Its deduced primary structure in the region spanning amino acid residues 1 to 147 of the protein exhibits 17 and 12 differences from LinA type 1 and LinA type 2, respectively. In addition, the residues GIHFAPS, present at the region spanning residues 148 to 154 in both LinA type 1 and LinA type 2, are deleted in LinA type 3.The activity of LinA type 3 for the metabolism of δ-HCH is several orders of magnitude higher than that of LinA type 1 or LinA type 2 and can be useful for improvement of the metabolism of δ-HCH.

scholarly journals Characterization of meprin, a membrane-bound metalloendopeptidase from mouse kidney

1987 ◽  
Vol 241 (1) ◽  
pp. 229-235 ◽  
Author(s):  
P E Butler ◽  
M J McKay ◽  
J S Bond
Keyword(s):  
Amino Acid ◽  
Degradation Products ◽  
Amino Acid Content ◽  
Peptide Bond ◽  
Mouse Kidney ◽  
Amino Acid Residues ◽  
Peptide Bonds ◽  
Membrane Bound ◽  
Terminal Amino

Meprin is an intrinsic protein of the brush border, a specialized plasma membrane, of the mouse kidney. It is a metalloendopeptidase that contains 1 mol of zinc and 3 mol of calcium per mol of the 85,000-Mr subunit. The enzyme is isolated, and active, as a tetramer. The behaviour of the enzyme on SDS/polyacrylamide gels in the presence and absence of beta-mercaptoethanol indicates that the subunits are of the same Mr (approx. 85,000) and held together by intersubunit S–S bridges. Eight S-carboxymethyl-L-cysteine residues were detected after reduction of the enzyme with beta-mercaptoethanol and carboxymethylation with iodoacetate. The enzyme is a glycoprotein and contains approx. 18% carbohydrate. Most of the carbohydrate is removed by endoglycosidase F, indicating that the sugar residues are N-linked. The isoelectric point of the enzyme is between pH 4 and 5, and the purified protein yields a pattern of evenly spaced bands in this range on isoelectric focusing. The peptide-bond specificity of the enzyme has been determined by using the oxidized B-chain of insulin as substrate. In all, 15 peptide degradation products were separated by h.p.l.c. and analysed for their amino acid content and N-terminal amino acid residue. The prevalent peptide-bond cleavages were between Gly20 and Glu21, Phe24 and Phe25 and between Phe25 and Tyr26. Other sites of cleavage were Leu6-Cysteic acid7, Ala14-Leu15, His10-Leu11, Leu17-Val18, Gly8-Ser9, Leu15-Tyr16, His5-Leu6. These results indicate that meprin has a preference for peptide bonds that are flanked by hydrophobic or neutral amino acid residues, but hydrolysis is not limited to these bonds. The ability of meprin to hydrolyse peptide bonds between small neutral and negatively charged amino acid residues distinguishes it from several other metalloendopeptidases.

Triazene derivatives of (1,x)-diazacycloalkanes — Part IX. Synthesis and characterization of a series of 1,4-di[2-aryl-1-diazenyl]-2,6-dimethylpiperazines

2010 ◽  
Vol 88 (4) ◽  
pp. 344-351 ◽  
Author(s):  
Naomi Hunter ◽  
Keith Vaughan
Keyword(s):  
Diazonium Salt ◽  
Mass Measurement ◽  
2D Nmr ◽  
Piperazine Ring ◽  
Ir And Nmr Spectroscopy ◽  
Methylene Groups ◽  
Type 3 ◽  
Derivatives Of

A series of 1,4-di-[2-aryl-1-diazenyl]-2,6-dimethylpiperazines (5a–5l), have been synthesized by the reaction of 2,6-dimethylpiperazine with 2 equiv. of the appropriate diazonium salt. The products have been characterized by IR and NMR spectroscopy, and the molecular composition has been verified by high-resolution EI mass spectrometry with accurate mass measurement of the molecular ion. The presence of stereocenters at C2 and C6 of the piperazine ring in the bis-triazene 5 creates two unique pairs of diastereotopic protons in the methylene groups at positions 3 and 5 of the piperazine ring, as evidenced by the complexity of the NMR spectra, which nevertheless can be fully assigned in most cases. The assignment of the proton and carbon signals in the 1,4-di-[2-aryl-1-diazenyl]-2,6-dimethylpiperazines has been aided by the use of 2D NMR HSQC spectroscopy. These results compare favorably with assignments of proton and carbon signals reported previously for triazenes of type 1 and bis-triazenes of type 3.

scholarly journals Drosophila melanogaster Mutated in its GBA1b Ortholog Recapitulates Neuronopathic Gaucher Disease

2019 ◽  
Vol 8 (9) ◽  
pp. 1420 ◽  
Author(s):  
Cabasso ◽  
Paul ◽  
Dorot ◽  
Maor ◽  
Krivoruk ◽  
...  
Keyword(s):  
Gaucher Disease ◽  
Disease Treatment ◽  
Therapeutic Modalities ◽  
Unfolded Protein ◽  
Terminal Amino ◽  
The Unfolded Protein Response ◽  
Protein Response ◽  
Type 3

Gaucher disease (GD) results from mutations in the GBA1 gene, which encodes lysosomal glucocerebrosidase (GCase). The large number of mutations known to date in the gene lead to a heterogeneous disorder, which is divided into a non-neuronopathic, type 1 GD, and two neurological, type 2 and type 3, forms. We studied the two fly GBA1 orthologs, GBA1a and GBA1b. Each contains a Minos element insertion, which truncates its coding sequence. In the GBA1am/m flies, which express a mutant protein, missing 33 C-terminal amino acids, there was no decrease in GCase activity or substrate accumulation. However, GBA1bm/m mutant flies presented a significant decrease in GCase activity with concomitant substrate accumulation, which included C14:1 glucosylceramide and C14:0 glucosylsphingosine. GBA1bm/m mutant flies showed activation of the Unfolded Protein Response (UPR) and presented inflammation and neuroinflammation that culminated in development of a neuronopathic disease. Treatment with ambroxol did not rescue GCase activity or reduce substrate accumulation; however, it ameliorated UPR, inflammation and neuroinflammation, and increased life span. Our results highlight the resemblance between the phenotype of the GBA1bm/m mutant fly and neuronopathic GD and underlie its relevance in further GD studies as well as a model to test possible therapeutic modalities.

scholarly journals Characterization of an infectious cDNA copy of the genome of a naturally occurring, avirulent coxsackievirus B3 clinical isolate

2005 ◽  
Vol 86 (1) ◽  
pp. 197-210 ◽  
Author(s):  
C.-K. Lee ◽  
K. Kono ◽  
E. Haas ◽  
K.-S. Kim ◽  
K. M. Drescher ◽  
...  
Keyword(s):  
Avirulent Strain ◽  
Cdna Copy ◽  
Naturally Occurring ◽  
Pathogenic Virus ◽  
Phenotype Comparison ◽  
Group B ◽  
Type 3 ◽  
Virus Strains

Group B coxsackieviruses (CVB) cause numerous diseases, including myocarditis, pancreatitis, aseptic meningitis and possibly type 1 diabetes. To date, infectious cDNA copies of CVB type 3 (CVB3) genomes have all been derived from pathogenic virus strains. An infectious cDNA copy of the well-characterized, non-pathogenic CVB3 strain GA genome was cloned in order to facilitate mapping of the CVB genes that influence expression of a virulence phenotype. Comparison of the sequence of the parental CVB3/GA population, derived by direct RT-PCR-mediated sequence analysis, to that of the infectious CVB3/GA progeny genome demonstrated that an authentic copy was cloned; numerous differences were observed in coding and non-coding sequences relative to other CVB3 strains. Progeny CVB3/GA replicated similarly to the parental strain in three different cell cultures and was avirulent when inoculated into mice, causing neither pancreatitis nor myocarditis. Inoculation of mice with CVB3/GA protected mice completely against myocarditis and pancreatitis induced by cardiovirulent CVB3 challenge. The secondary structure predicted for the CVB3/GA domain II, a region within the 5′ non-translated region that is implicated as a key site affecting the expression of a cardiovirulent phenotype, differs from those predicted for cardiovirulent and pancreovirulent CVB3 strains. This is the first report characterizing a cloned CVB3 genome from an avirulent strain.

scholarly journals Construction and Characterization of E3-Deleted Bovine Adenovirus Type 3 Expressing Full-Length and Truncated Form of Bovine Herpesvirus Type 1 Glycoprotein gD

Virology ◽  
1998 ◽  
Vol 250 (1) ◽  
pp. 220-229 ◽  
Author(s):  
Alexandre N. Zakhartchouk ◽  
P.Seshidhar Reddy ◽  
Mohit Baxi ◽  
Maria E. Baca-Estrada ◽  
Majid Mehtali ◽  
...  
Keyword(s):  
Bovine Herpesvirus ◽  
Adenovirus Type ◽  
Full Length ◽  
Truncated Form ◽  
Herpesvirus Type ◽  
Bovine Herpesvirus Type 1 ◽  
Bovine Adenovirus Type 3 ◽  
Type 3

Synthesis, structure and characterization of five new organically templated metal sulfates with 2-aminopyridinium

2016 ◽  
Vol 72 (5) ◽  
pp. 432-441 ◽  
Author(s):  
Tamara J. Bednarchuk ◽  
Vasyl Kinzhybalo ◽  
Adam Pietraszko
Keyword(s):  
Materials Science ◽  
Three Dimensional ◽  
X Ray Diffraction ◽  
Hydrogen Bond Formation ◽  
Unit Cells ◽  
Science Community ◽  
Type 3

The chemistry of organically templated metal sulfates has attracted interest from the materials science community and the development of synthetic strategies for the preparation of organic–inorganic hybrid materials with novel structures and special properties is of current interest. Sulfur–oxygen–metal linkages provide the possibility of using sulfate tetrahedra as building units to form new solid-state materials. A series of novel organically templated metal sulfates of 2-aminopyridinium (2ap) with aluminium(III), cobalt(II), magnesium(II), nickel(II) and zinc(II) were obtained from the respective aqueous solutions and studied by single-crystal X-ray diffraction. The compounds crystallize in centrosymmetric triclinic unit cells in three structure types: type 1 for 2-aminopyridinium hexaaquaaluminium(III) bis(sulfate) tetrahydrate, (C5H7N2)[Al(H2O)6](SO4)2·4H2O, (I); type 2 for bis(2-aminopyridinium) tris[hexaaquacobalt(II)] tetrakis(sulfate) dihydrate, (C5H7N2)2[Co(H2O)6]3(SO4)4·2H2O, (II), and bis(2-aminopyridinium) tris[hexaaquamagnesium(II)] tetrakis(sulfate) dihydrate, (C5H7N2)2[Mg(H2O)6]3(SO4)4·2H2O, (III); and type 3 for bis(2-aminopyridinium) hexaaquanickel(II) bis(sulfate), (C5H7N2)2[Ni(H2O)6](SO4)2, (IV), and bis(2-aminopyridinium) hexaaquazinc(II) bis(sulfate), (C5H7N2)2[Zn(H2O)6](SO4)2, (V). The templating role of the 2ap cation in all of the reported crystalline substances is governed by the formation of characteristic charge-assisted hydrogen-bonded pairs with sulfate anions and the presence of π–π interactions between the cations. Additionally, both coordinated and uncoordinated water molecules are involved in hydrogen-bond formation. As a consequence, extensive three-dimensional hydrogen-bonding patterns are formed in the reported crystal structures.

Altered expression of renal AQPs and Na+transporters in rats with Lithium-induced NDI

2000 ◽  
Vol 279 (3) ◽  
pp. F552-F564 ◽  
Author(s):  
Tae-Hwan Kwon ◽  
Ulla H. Laursen ◽  
David Marples ◽  
Arvid B. Maunsbach ◽  
Mark A. Knepper ◽  
...  
Keyword(s):  
Nephrogenic Diabetes Insipidus ◽  
Loop Of Henle ◽  
Low Doses ◽  
Altered Expression ◽  
Aquaporin 1 ◽  
High Doses ◽  
Type 3 ◽  
And Control

Lithium (Li) treatment is often associated with nephrogenic diabetes insipidus (NDI). The changes in whole kidney expression of aquaporin-1 (AQP1), -2, and -3 as well as Na-K-ATPase, type 3 Na/H exchanger (NHE3), type 2 Na-Pi cotransporter (NaPi-2), type 1 bumetanide-sensitive Na-K-2Cl cotransporter (BSC-1), and thiazide-sensitive Na-Cl cotransporter (TSC) were examined in rats treated with Li orally for 4 wk: protocol 1, high doses of Li (high Na+ intake), and protocol 2, low doses of Li (identical food and normal Na+ intake in Li-treated and control rats). Both protocols resulted in severe polyuria. Semiquantitative immunoblotting revealed that whole kidney abundance of AQP2 was dramatically reduced to 6% ( protocol 1) and 27% ( protocol 2) of control levels. In contrast, the abundance of AQP1 was not decreased. Immunoelectron microscopy confirmed the dramatic downregulation of AQP2 and AQP3, whereas AQP4 labeling was not reduced. Li-treated rats had a marked increase in urinary Na+ excretion in both protocols. However, the expression of several major Na+ transporters in the proximal tubule, loop of Henle, and distal convoluted tubule was unchanged in protocol 2, whereas in protocol 1significantly increased NHE3 and BSC-1 expression or reduced NaPi-2 expression was associated with chronic Li treatment. In conclusion, severe downregulation of AQP2 and AQP3 appears to be important for the development of Li-induced polyuria. In contrast, the increased or unchanged expression of NHE3, BSC-1, Na-K-ATPase, and TSC indicates that these Na+ transporters do not participate in the development of Li-induced polyuria.

scholarly journals Utility of Japan Narrow Band Imaging Expert Team Classification Using Narrow Band Imaging for Evaluation of Colonic Polyps

2020 ◽  
Vol 11 (02) ◽  
pp. 138-145
Author(s):  
Dipak S. Ahire ◽  
Pravin M. Rathi ◽  
Niranjan H. Banka ◽  
Parth K. Shah
Keyword(s):  
Predictive Value ◽  
Narrow Band ◽  
Narrow Band Imaging ◽  
Colonic Polyps ◽  
Current Evidence ◽  
Low Grade ◽  
Expert Team ◽  
Type 3

Abstract Background Narrow band imaging (NBI) is an advanced endoscopic imaging technique that enhances visualization of the mucosal surface and is used as a screening tool for colonic polyps. Its usefulness is currently explored to a lesser extent in India. So, we assessed the utility of Japan NBI Expert Team (JNET) classification for characterization of colorectal polyps. Methods A prospective observational study was performed from January 2018 to June 2019 of patients undergoing colonoscopy at a tertiary care hospital. NBI image of polyps was captured followed by either polypectomy/biopsy. Histopathology results were correlated with the pattern revealed by NBI on polyps using the JNET classification. Results A total of 80 patients, 61(76.25%) male with a mean (standard deviation [SD]) age of 58.41 ± 14.59 years were included. Out of the 90 lesions, 23 (25.5%) had type-1 pattern, 45 (50%) had 2A, 13 (14.4%) had 2B, and 9 (10%) had type-3 pattern. On histopathology, majority 51 (59.3%) were found to be adenomatous with low-grade intramucosal neoplasia. When correlating our results with JNET category type 1 and hyperplastic polyps, the sensitivity was 90%, specificity was 97%, negative predictive value was 97%, positive predictive value was 90%, and diagnostic accuracy was 96%. Correlating type 2A and low-grade intramucosal neoplasia had results of 78, 87, 76, 90, and 82%, respectively. Correlating type 2B and high-grade intramucosal neoplasia had results of 83, 90, 99, 38, and 90%, respectively. Correlating type 3 and deep submucosal cancer had results of 88, 98, 99, 78, and 97%, respectively. Conclusion NBI shows excellent probability to exclude carcinoma possibilities based on the changes in colonic mucosal features. Owing to slightly lower sensitivity for type 2B, it needs additional investigation using pit pattern diagnosis. We demonstrated the high-diagnostic performance of NBI in making an accurate diagnosis of early colorectal cancers in colonoscopy. Further refinement in the NBI technology might add to the current evidence for characterization of polyps.

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