Unformatted text preview: (D) The organelles in the pellet fro m step (C) were separated from one another by sedimentation equilibrium in a sucrose densit y gradient. Each of the above fractions was treated with detergent and then co mbined with ant ibodies against carboxypept idase Y, a vacuolar yeast protein. The immunoprecipitates fro m each fract ion were counted in a gamma counter and the cpm obtained for the immunoprecipitates fro m each fract ion at each time interval are listed in the table below. __________________CHASE TIME______________________ CELL FRACTION 5 min 10 min Cell medium Golgi Microsomes Mitochondria Vacuoles 0 100 24,800 0 0 0 22,000 100 0 0 15 min 0 2,114 27 0 23,500 25 min 0 53 0 0 24,857 Answer the fo llowing quest ions: a) How long does it take newly synt hesized carboxypept idase Y to reach the vacuo les? On the basis of the above data, name the organelles, in order, through which carboxypept idase Y passes from its site of synt hesis on the ribosomes en route to the vacuo les. b) Why was sucrose included in steps B, C and D above? 10. A patch of membrane containing badrenergic receptors and other relevant membrane proteins was pulled fro m a cell using a glass pipette, similar to the process used in a patch clamping technique. As shown in the diagram below, the external surface of the membrane is in contact with the solution in the pipette and the cytoplasmic surface o f the membrane is immersed in a small well. The well contains + + buffer that is equivalent to the salt composit ion (Mg +, Ca +, etc.) of the cytosol and contains ATP, but does not contain other small mo lecules such as other nucleotides. pipette intact cell De mo
pipette well outer solution inner solution ...
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- Spring '08
- carboxypept idase, Golgi Microsomes Mitochondria, pellet fro, TIME______________________ CELL FRACTION, relevant membrane proteins, vacuolar yeast protein.