Isolation of renin granules from rat kidney cortex by isotonic or hyperosmotic metrizamide-sucrose gradients.
The crude renin granules isolated from rat kidney cortex were purified using iso-osmotic metrizamide-sucrose gradients having osmolalities ranging from 300 mOsm/kg (= isotonic) to 1700 mOsm/kg. The density gradients were centrifuged at 116,000 g (maximum) for 60 min. 1. Working at 22 degrees-25 degrees C gave similar results as working at 0 degrees-4 degrees C. Hence further experiments were performed at room temperature. 2. The density of renin granules, mitochondria and lysosomes was a linear positive function of the log of osmolality. 3. It was not possible to separate renin granules from mitochondria at 300 mOsm/kg since both the organelles equilibrated at 1.154 kg/l. A reproducible separation was achieved at 850 mOsm/kg or higher but then the major fraction of lysosomes was superimposed on renin granules. Microsomes were always lighter than renin granules. 4. As compared with the total homogenates, renin/protein ratio was increased six-fold renin/malic acid dehydrogenase ratio 21-fold and renin/beta-glucuronidase ratio 3.5-fold. 5. Finally it was demonstrated that renin granules purified at high osmolalities tend to lyse when transferred into an isotonic medium.
D000818:Animals; D002458:Cell Fractionation / Q000379:methods; D002499:Centrifugation, Density Gradient / Q000379:methods; D007552:Isotonic Solutions; D007672:Kidney Cortex / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology; D008291:Malate Dehydrogenase / Q000302:isolation & purification; D008793:Metrizamide; D008928:Mitochondria / Q000201:enzymology; D009994:Osmolar Concentration; D051381:Rats; D012083:Renin / Q000302:isolation & purification*; D013395:Sucrose; D013696:Temperature
D007552:Isotonic Solutions; D013395:Sucrose; D008291:Malate Dehydrogenase; D012083:Renin; D008793:Metrizamide