Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.26.9 (ribonuclease)
 6,589 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The beta-adrenergic receptor kinase (beta ARK) specifically phosphorylates the agonist-occupied form of the beta-adrenergic and related G protein-coupled receptors. Structural features of this enzyme have been elucidated recently by the isolation of a cDNA that encodes bovine beta ARK. Utilizing a catalytic domain fragment of the beta ARK cDNA to screen a bovine brain cDNA library we have isolated a clone encoding a beta ARK-related enzyme which we have termed beta ARK2. Overall, this enzyme has 85% amino acid identity with beta ARK, with the protein kinase catalytic domain having 95% identity. The ability of beta ARK2 to phosphorylate various substrates was studied after expression in COS 7 cells. Although beta ARK2 is essentially equiactive with beta ARK in phosphorylating an acid-rich synthetic model peptide it was only approximately 50% as active when the substrate was the agonist-occupied beta 2-adrenergic receptor and only approximately 20% as active toward light-bleached rhodopsin. As with beta ARK, phosphorylation of the receptor substrates by beta ARK2 was completely stimulus dependent. RNA blot analysis with selected bovine tissues reveals an mRNA of 8 kilobases with a distribution similar to that of beta ARK. More detailed RNA analysis using a ribonuclease protection assay in various rat tissues suggests that the beta ARK2 message is present at much lower levels (typically 10-20%) than the beta ARK message. In the rat the beta ARK2 mRNA is localized predominantly in neuronal tissues although low levels are also observed in various peripheral tissues. The beta ARK2 gene has been localized to a region of mouse chromosome 5 whereas the beta ARK gene is localized on mouse chromosome 19. These data suggest the existence of a "family" of receptor kinases which may serve broadly to regulate receptor function.
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PMID:Cloning, expression, and chromosomal localization of beta-adrenergic receptor kinase 2. A new member of the receptor kinase family. 186 33

The principal mechanism of homologous desensitization of the beta-adrenergic receptor (beta2AR) is phosphorylation of the receptor by the betaAR kinase (betaARK) or other closely related G protein-coupled receptor kinases (GRKs). However, within a single organ such as the lung where many cell types express the receptor, the presence or extent of beta2AR desensitization in different cells has been noted to be highly variable. We hypothesized that such variability in desensitization is due to significant cell-type differences in betaARK expression and/or function. To approach this, in situ hybridization was carried out in the lung and indeed revealed heterogeneity in betaARK gene expression. Quantitative studies using ribonuclease protection assays with cell lines revealed that the level of betaARK mRNA in airway smooth muscle cells was approximately 20% of that in bronchial epithelial cells and approximately 11% of that in mast cells (6.65 +/- 0.96 versus 32.6 +/- 4.0 and 60.7 +/- 1.5 relative units, respectively, p < 0. 001). betaARK2 gene expression was not detected in any of these cells. At the protein level, betaARK expression in airway smooth muscle cells was nearly undetectable, being approximately 10-fold less than that expressed on mast cells. The activities of the GRKs in cell extracts were assessed in vitro by quantitating their ability to phosphorylate rhodopsin in the presence of light. Consistent with the gene and protein expression results, a marked discrepancy in activities was observed between extracts derived from mast cells (90.7 +/- 0.5 relative units) as compared to airway smooth muscle cells (9.28 +/- 0.6 relative units, p < 0.001). In contrast, the activities of protein kinase A (the other kinase that phosphorylates beta2AR) in these extracts were not different. We predicted, then, that airway smooth muscle beta2AR would undergo minimal short-term (5 min) agonist-promoted desensitization as compared to the beta2AR expressed on mast cells. Mast cell cAMP reached maximal levels after 90 s and did not further increase over time, indicative of receptor desensitization in this cell. In contrast, cAMP levels of airway smooth muscle cells did not plateau, increasing at a rate of 103 +/- 9% per min, consistent with little desensitization over the study period. We conclude that there is significant cell-type variation in expression of betaARK and that such variation is directly related to the extent of short-term agonist-promoted desensitization of the beta2AR.
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PMID:Heterogeneity in beta-adrenergic receptor kinase expression in the lung accounts for cell-specific desensitization of the beta2-adrenergic receptor. 905 32

We have used the most advanced programs currently available to construct the first three-domain structure of the human thyrotropin receptor (TSHR) using comparative modeling. The model consists of a leucine-rich domain (LRD; amino acids 36-281; porcine ribonuclease inhibitor used as a template for modeling), a cleavage domain (CD; amino acids 282-409; tissue inhibitor of matrix metalloproteinases 2 as template) and transmembrane domain (TMD amino acids 410-699; bovine rhodopsin as template). Models of human, porcine, and bovine TSH were also constructed (human chorionic gonadotropin [hCG] and human follicle stimulating hormone [hFSH] as templates). The LRD has a characteristic horseshoe shape with 10 tandem homologous repeats. The CD consists of beta-barrel and alpha helix structures (OB-like fold) with two disulfide bridges and the structure around these disulfide bridges remains stable after cleavage. The TMD presents the typical seven membrane-spanning helices. The TSH, LRD, CD, and TMD models were brought together in an extensive series of docking experiments. Known features of the TSH-TSHR interaction were used for selection of appropriate complexes that were then validated using a different set of experimental data. A similar approach was used to build a model of a complex between the TSHR and a monoclonal TSHR antibody with weak thyroid stimulating activity. Human thyrotropin (hTSH) alpha chains were found to make contact with many amino acids on the LRD surface and CD surface whereas no interaction between the beta chains and the CD were found. The higher affinity of bovine thyrotropin (bTSH) and porcine thyrotropin (pTSH) (relative to hTSH) for the TSHR is explained well by the models in terms of charge-charge interactions between their alpha chains and the receptor. Experimental observations showing increased sensitivity of the TSHR to hCG after mutation of TSHR Lys209 to Glu are explained well by our model. Furthermore, several mutations in the TMD that are associated with increased TSHR basal activity are predicted from our model to be caused by the formation of new interactions that stabilize the activated form of the TMD.
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PMID:Analysis of the thyrotropin receptor-thyrotropin interaction by comparative modeling. 1617 67