76 and 14 kDa polypeptides, two major components released from amphibian urinary bladder epithelium. Purification and characterization.
Exocytotic processes play a major role in the hormonal control of water permeability in the amphibian urinary bladder. Different treatments such as antidiuretic hormone (ADH) stimulation, incubation with phorbol ester or mild detergent and mechanical stretch of the bladder, consistently induce a liberation of two major polypeptides of 76 and 14 kDa molecular mass into the luminal medium. Each of these polypeptides represents 3 to 5% of the total protein of epithelial cell homogenates and 20 to 50% of the released material. Proportions of released 76 kDa polypeptide in urinary bladders of toads (Bufo marinus) and frogs (Rana esculenta) were similar but, in the frog extracts, two bands ("doublet") were resolved at the level of 76 kDa. In high performance liquid chromatography (HPLC), using gel filtration and ion exchange chromatography, the frog 76 kDa protein was resolved into two polypeptides of 80,000 to 100,000 and 60,000 to 80,000 daltons while the 14 kDa protein included two polypeptides, each with a molecular mass of approximately 14,000 daltons. Isoelectric focusing of the material released during a mechanical stretch of the tissue ("stretch extract") or of isolated purified proteins from the frog urinary bladder showed that the 14 kDa polypeptides were resolved in two major groups of polypeptides, one in the range of pH 7.4 to 7.8, the other at pH 5.6. The lower band of the 76 kDa doublet also comprised some diffuse bands (5.0 less than pI less than 5.2) while the other polypeptide of the doublet presented a sharp band at pH 6.2.(ABSTRACT TRUNCATED AT 250 WORDS)
Dassouli
A
A
Unité de Recherches sur la Pathologie Rénale et Vasculaire, INSERM U 28, Hôpital Broussais, Paris/France.
Chevalier
J
J
Ripoche
P
P
D000818:Animals; D002024:Bufo marinus; D007106:Immune Sera; D008970:Molecular Weight; D010703:Phorbol Esters / Q000494:pharmacology*; D011506:Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D011893:Rana esculenta; D001743:Urinary Bladder / Q000187:drug effects / Q000378:metabolism*; D014667:Vasopressins / Q000494:pharmacology*
D007106:Immune Sera; D010703:Phorbol Esters; D011506:Proteins; D014667:Vasopressins