scholarly journals Identification and characterization of a novel A-Kinase-anchoring protein (AKAP120) from rabbit gastric parietal cells

1997 ◽  
Vol 322 (3) ◽  
pp. 801-808 ◽  
Author(s):  
Daniel T. DRANSFIELD ◽  
Jennifer L. YEH ◽  
Alvin J. BRADFORD ◽  
James R. GOLDENRING
Keyword(s):  
Amino Acids ◽  
Gastric Fundus ◽  
Parietal Cells ◽  
Regulatory Subunit ◽  
Binding Region ◽  
Rnase Protection ◽  
Gastric Parietal Cell ◽  
High Affinity ◽  
Gastric Parietal Cells ◽  
Recombinant Fragments

The type-II cAMP-dependent protein kinase (A-Kinase) partitions primarily into the particulate fraction in gastric parietal cells. Localization of this kinase to particular subcellular domains is mediated through the binding of the regulatory subunit (RII) dimer to A-Kinase-anchoring proteins (AKAPs). Using a [32P]RII overlay assay, we have screened a rabbit gastric parietal cell cDNA library and have isolated a single RII-binding protein clone. Sequence analysis revealed an open reading frame coding for 1022 amino acids (AKAP120). Recombinant fragments of the full-length clone were prepared and the RII-binding region mapped to an area between amino acids 489 and 549. This area contained a putative α-helical RII-binding region between amino acids 503 and 516. Incubation of [32P]RII with a synthetic peptide of AKAP120-(489–522) completely inhibited the binding of [32P]RII to the recombinant AKAP120 fragments that demonstrated RII binding. In vitroRII -binding affinity studies indicated a high-affinity interaction between AKAP120 and RII with a Kapp between 50 and 120 nM for the three recombinant fragments that bound [32P]RII. RNase-protection analysis revealed that AKAP120 is a widely distributed protein, with the highest levels of mRNA observed in gastric fundus. The presence of this novel high-affinity AKAP in gastric parietal cells suggests that it may regulate RII subcellular sequestration in this cell type.

Gastric parietal cell H(+)-K(+)-ATPase microsomes are associated with isoforms of ankyrin and spectrin

1993 ◽  
Vol 264 (1) ◽  
pp. C63-C70 ◽  
Author(s):  
P. R. Smith ◽  
A. L. Bradford ◽  
E. H. Joe ◽  
K. J. Angelides ◽  
D. J. Benos ◽  
...  
Keyword(s):  
Apical Membrane ◽  
Immunoblot Analysis ◽  
Cytoskeletal Proteins ◽  
Parietal Cells ◽  
Apical Surface ◽  
Gastric Parietal Cell ◽  
Apical Domain ◽  
Hcl Secretion ◽  
Gastric Parietal Cells ◽  
Stimulation Of

Stimulation of HCl secretion by gastric parietal cells requires the fusion of cytoplasmic H(+)-K(+)-ATPase-bearing tubulovesicles with the apical membrane. This insertion of membrane results in a dramatic increase in apical surface area through the formation of microvilli. To elucidate the elements that may stabilize the newly inserted H(+)-K(+)-ATPase within the apical membrane, we searched for specific cytoskeletal proteins associating with the gastric enzyme. We document by immunoblot analysis that ankyrin, spectrin, and actin copurify with H(+)-K(+)-ATPase microsomes prepared from gastric parietal cells. Coprecipitation of 125I-labeled native erythrocyte ankyrin with the H(+)-K(+)-ATPase from gastric microsomes using anti-H(+)-K(+)-ATPase antibodies suggests that ankyrin associates with the H(+)-K(+)-ATPase. Indirect immunofluorescence and confocal microscopy show that ankyrin and H(+)-K(+)-ATPase cosegregate within resting and secreting parietal cells. Taken together, these data suggest that the association of the gastric H(+)-K(+)-ATPase with spectrin and actin is mediated by ankyrin and that this interaction contributes to the maintenance of the polarized distribution of the enzyme to the apical domain of gastric parietal cells during acid secretion.

scholarly journals Anti-Helicobacter Pylori, Anti-Thyroid Peroxidase, Anti-Thyroglobulin and Anti-Gastric Parietal Cells Antibodies in Czech Population

2008 ◽  
pp. S135-S141
Author(s):  
I Šterzl ◽  
P Hrdá ◽  
P Matucha ◽  
J Čeřovská ◽  
V Zamrazil
Keyword(s):  
Helicobacter Pylori ◽  
Thyroid Autoimmunity ◽  
Parietal Cells ◽  
Thyroid Peroxidase ◽  
Gastric Parietal Cell ◽  
Czech Population ◽  
Study Support ◽  
H Pylori ◽  
Gastric Parietal Cells ◽  
Age And Sex

Autoimmune thyropathies are frequently linked to many infections, such as Helicobacter pylori, which are also supposed to play a role in their pathogenesis. The aim of this study was to evaluate the relationships between thyroid and gastric autoimmunity and H. pylori infection on a large sample of Czech population (n=1621) by monitoring the autoantibodies against thyroglobulin (anti-Tg) and thyroid peroxidase (anti-TPO) and gastric parietal cell (anti-GPC, representing thyrogastric syndrome) in correlation with antibodies against Helicobacter pylori (anti-H. pylori) of classes IgG and IgA. The interrelation between autoantibodies and H. pylori antibodies was assessed by H. pylori seropositivity. In H. pylori seropositive persons as compared to seronegative irrespective of age and sex, a higher occurrence of anti-TPO (10.4 % vs. 5.8 %, p=0.001) and anti-GPC (6.1 % vs. 1.7 %, p<0.001) was found. Differences in anti-TPO occurrence were significant in both men (7.0 % vs. 3.3 %, p=0.03) and women (12.7 % vs. 8.0 %, p=0.02), differences in anti-GPC occurrence were significant only in women (7.2 % vs. 1.7 %, p<0.001). Results of this study support the idea of a connection between infection of H. pylori and the occurrence of anti-TPO autoantibodies representing thyroid autoimmunity and gastric parietal cells autoantibodies representing the thyrogastric syndrome.

La3+ and pH sensitivity of Ca2+ entry and intracellular store filling in gastric parietal cells

1995 ◽  
Vol 269 (5) ◽  
pp. G770-G778 ◽  
Author(s):  
P. A. Negulescu ◽  
T. E. Machen
Keyword(s):  
Positive Feedback ◽  
Low Ph ◽  
Parietal Cells ◽  
Additional Increase ◽  
Cholinergic Agonist ◽  
Intracellular Store ◽  
Entry Pathway ◽  
Signaling Function ◽  
Previously Treated ◽  
Gastric Parietal Cells

The fluorescent Ca2+ indicator fura 2 was used to measure cytosolic free [Ca2+] ([Ca2+]i) in order to obtain information about relative rates of Ca2+ influx into parietal cells during treatment with carbachol (a cholinergic agonist) or thapsigargin (TG, a Ca(2+)-mobilizing agent) or during reloading of the internal Ca2+ stores. In Ca(2+)-containing solutions, carbachol-, TG-, and reloading-stimulated Ca2+ entry exhibited nearly identical sensitivity to La3+ [inhibition constant (Ki) approximately 10 microM] or low pH (pKi approximately 7.0). In experiments in which carbachol and TG were used, there was no additional increase in [Ca2+]i when TG was added to carbachol-treated cells or when carbachol was added to cells previously treated with TG. Thus it is likely that a single Ca2+ entry pathway serves a signaling function as well as a role in refilling the Ca2+ store during reloading. Because the Ca2+ pathway is exquisitely sensitive to pH and serosal pH increases during stimulant-induced H+ secretion (which is activated by increases in [Ca2+]i), this mechanism will exert positive feedback on parietal cells in the intact stomach. When parietal cells were pretreated with carbachol in Ca(2+)-free solutions, reloading was independent of pH and La3+, suggesting that Ca(2+)-containing solutions should be used to determine the properties of the influx pathway.

Molecular docking studies of tea (Thea sinensis Linn.) polyphenols inhibition pattern with Rat P-glycoprotein

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Babar Ali ◽  
Qazi Mohammad Sajid Jamal ◽  
Showkat R. Mir ◽  
Saiba Shams ◽  
Mohammad Amjad Kamal
Keyword(s):  
Amino Acids ◽  
Molecular Docking ◽  
Binding Energy ◽  
Oral Administration ◽  
Docking Studies ◽  
Vital Role ◽  
Glycoprotein P ◽  
High Affinity ◽  
P Glycoprotein ◽  
Inhibition Pattern

AbstractSince 3000 B.C., evergreen plant Thea sinensis (Theaceae) is used both as a social and medicinal beverage. Leaves of T. sinensis contain amino acids, vitamins, caffeine, polysaccharides and polyphenols. Most of the natural medicinal actions of tea are due to the availability and abundance of polyphenols mainly catechins. It has also been stated that some catechins were absorbed more rapidly than other compounds after the oral administration of tea and could increase the bio-enhancing activities of anticancer drugs by inhibiting P-glycoprotein (P-gp). The results of the molecular docking showed that polyphenols bind easily to the active P-gp site. All compounds exhibited fluctuating binding affinity ranged from −11.67 to −8.36 kcal/mol. Observed binding energy required for theaflavin to bind to P-gp was lowest (−11.67 kcal/mol). The obtained data that supports all the selected polyphenols inhibited P-gp and therefore may enhance the bioavailability of drugs. This study may play a vital role in finding hotspots in P-gp and eventually may be proved useful in designing compounds with high affinity and specificity to the protein.

scholarly journals Receptor-operated Ca2+ channels in gastric parietal cells: gastrin and carbachol induce Ca2+ influx in depleting intracellular Ca2+ stores

1993 ◽  
Vol 289 (1) ◽  
pp. 117-124 ◽  
Author(s):  
S Roche ◽  
J P Bali ◽  
R Magous
Keyword(s):  
Steady State ◽  
Receptor Activation ◽  
Parietal Cells ◽  
The Other ◽  
Bivalent Cation ◽  
Gtp Binding ◽  
Gastric Parietal Cells ◽  
Type Receptor ◽  
Ca2 Channels ◽  
Stimulation Of

The mechanism whereby gastrin-type receptor and muscarinic M3-type receptor regulate free intracellular Ca2+ concentration ([Ca2+]i) was studied in rabbit gastric parietal cells stimulated by either gastrin or carbachol. Both agonists induced a biphasic [Ca2+]i response: a transient [Ca2+]i rise, followed by a sustained steady state depending on extracellular Ca2+. Gastrin and carbachol also caused a rapid and transient increase in Mn2+ influx (a tracer for bivalent-cation entry). Pre-stimulation of cells with one agonist drastically decreased both [Ca2+]i increase and Mn2+ influx induced by the other. Neither diltiazem nor pertussistoxin treatment had any effect on agonist-stimulated Mn2+ entry. Thapsigargin, a Ca(2+)-pump inhibitor, induced a biphasic [Ca2+]i increase, and enhanced the rate of Mn2+ entry. Preincubation of cells with thapsigargin inhibits the [Ca2+]i increase as well as Mn2+ entry stimulated by gastrin or by carbachol. Thapsigargin induced a weak but significant increase in Ins(1,4,5)P3 content, but this agent had no effect on the agonist-evoked Ins(1,4,5)P3 response. In permeabilized parietal cells, Ins(1,4,5)P3 and caffeine caused an immediate Ca2+ release from intracellular pools, followed by a reloading of Ca2+ pools which can be prevented in the presence of thapsigargin. We conclude that (i) gastrin and carbachol mobilize common Ca2+ intracellular stores, (ii) Ca2+ permeability secondary to receptor activation involves neither a voltage-sensitive Ca2+ channel nor a GTP-binding protein from the G1 family, and (iii) agonists regulate common Ca2+ channels in depleting intracellular Ca2+ stores.

scholarly journals Human Rev1 relies on insert-2 to promote selective binding and accurate replication of stabilized G-quadruplex motifs

2021 ◽  
Author(s):  
Amit Ketkar ◽  
Lane Smith ◽  
Callie Johnson ◽  
Alyssa Richey ◽  
Makayla Berry ◽  
...  
Keyword(s):  
Amino Acids ◽  
Mutation Frequency ◽  
Control Sequence ◽  
Wild Type ◽  
Binding Properties ◽  
High Affinity ◽  
G4 Dna ◽  
G Quadruplex ◽  
Forward Mutagenesis

Abstract We previously reported that human Rev1 (hRev1) bound to a parallel-stranded G-quadruplex (G4) from the c-MYC promoter with high affinity. We have extended those results to include other G4 motifs, finding that hRev1 exhibited stronger affinity for parallel-stranded G4 than either anti-parallel or hybrid folds. Amino acids in the αE helix of insert-2 were identified as being important for G4 binding. Mutating E466 and Y470 to alanine selectively perturbed G4 binding affinity. The E466K mutant restored wild-type G4 binding properties. Using a forward mutagenesis assay, we discovered that loss of hRev1 increased G4 mutation frequency &gt;200-fold compared to the control sequence. Base substitutions and deletions occurred around and within the G4 motif. Pyridostatin (PDS) exacerbated this effect, as the mutation frequency increased &gt;700-fold over control and deletions upstream of the G4 site more than doubled. Mutagenic replication of G4 DNA (±PDS) was partially rescued by wild-type and E466K hRev1. The E466A or Y470A mutants failed to suppress the PDS-induced increase in G4 mutation frequency. These findings have implications for the role of insert-2, a motif conserved in vertebrates but not yeast or plants, in Rev1-mediated suppression of mutagenesis during G4 replication.

Cloning and characterization of a human brain Na + -independent transporter for small neutral amino acids that transports d -serine with high affinity

2000 ◽  
Vol 287 (3) ◽  
pp. 231-235 ◽  
Author(s):  
Jun Nakauchi ◽  
Hirotaka Matsuo ◽  
Do Kyung Kim ◽  
Akiteru Goto ◽  
Arthit Chairoungdua ◽  
...  
Keyword(s):  
Amino Acids ◽  
Human Brain ◽  
Neutral Amino Acids ◽  
High Affinity
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