Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.5.1.52 (PNGase F)
 1,527 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We previously demonstrated that the AT2 receptor is a glycoprotein containing N-linked oligosaccharide side chains and that the marked disparity between the sizes of AT2 receptors from different tissues was related to different degrees of N-glycosylation. In the present study, we used an inhibitor of N-glycosylation, tunicamycin, as well as an endoglycosidase, glycopeptidase-F, to examine the contribution of carbohydrate moieties to the ligand-binding properties, cell-surface expression and apparent molecular mass of AT2 receptors of rat pheochromocytoma cells (PC-12 cells). Photoaffinity labelling of cell-surface AT2 receptors revealed that PC-12 cells grown in the presence of tunicamycin expressed, in addition to the previously described 140 kDa receptor, lower-molecular-mass receptors of 63 kDa, 47 kDa and 32 kDa. Lectin affinity chromatography revealed that the 63 kDa and the 47 kDa receptors are partially glycosylated and that the 32 kDa receptor is completely deglycosylated. Competitive binding experiments were carried out on tunicamycin-treated cells that expressed predominantly the 63 kDa or the 47 kDa receptors. Both receptor forms exhibited a high affinity for angiotensin II, although a slight decrease (of about 2-fold) was consistently observed on tunicamycin-treated cells as compared with control cells. Endoglycosidase digestion of AT2 receptors of PC-12 cells also yielded smaller receptor forms of 47 kDa and 32 kDa. Similarly, angiotensin II showed a high but slightly decreased binding affinity (of about 2-fold) for deglycosylated membranes as compared with control membranes. In conclusion, the stepwise action of tunicamycin suggests the presence of at least three N-linked oligosaccharide side chains on the AT2 receptor of PC-12 cells. These oligosaccharide side chains have a minor contribution to the affinity of the receptor. Interestingly, glycosylation is not an essential requirement for the expression of AT2 receptor at the surface of PC-12 cells.
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PMID:Analysis of the role of N-glycosylation in cell-surface expression and binding properties of angiotensin II type-2 receptor of rat pheochromocytoma cells. 854 98

Angiotensin II (AngII) binding sites were characterized on rat pheochromocytoma cells (PC-12) which are known to express exclusively the type-2 (AT2) AngII receptor. Interestingly, we found that, on confluent PC-12 cells, only partial inhibition of 125I-AngII binding was achieved when cells were incubated with a saturating concentration of PD-123 319 (an AT2 selective ligand) suggesting the presence of an atypical binding site. In binding experiments, AngII exhibited high affinity for this atypical binding site with a dissociation constant (Kd) of 16 nM. Moreover, bacitracin potently inhibited PD-123 319-resistant 125I-AngII binding with an IC50 half-maximal inhibitory concentration of 44 microM. Enzyme immunoassay revealed that the cells were contaminated with Mycoplasma hyorhynis. Contaminated PC-12 cells were photolabeled with 125I-[p-benzoylPhe1]AngII and covalently labeled proteins were subjected to polyacrylamide gel electrophoresis followed by autoradiography. Under these conditions, two distinct labeled species of 140 kilodaltons (kDa) and 95 kDa were detected. Deglycosylation of the 140 kDa-labeled AT2 receptor with glycopeptidase-F (PNGase-F) resulted in a 35 kDa protein whereas the 95 kDa band was not affected by digestion with the endoglycosidase. Thus, our results show that the AngII binding site on M. hyorhynis is structurally distinct from mammalian AT1 and AT2 receptors.
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PMID:The angiotensin II binding site on Mycoplasma hyorhynis is structurally distinct from mammalian AT1 and AT2 receptors. 953 71