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Studies on isolated subcellular components of cat pancreas

II. A Ca++-dependent interaction between membranes and zymogen granules of cat pancreas

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Summary

A membrane and zymogen granule fraction of cat pancreas has been purified on an exponential ficoll-sucrose gradient in a zonal rotor. A Ca++-dependent interaction between the membranes labelled with125I or14C-p-chloromercuribenzoate or N-ethyl(2,3-14C)maleimide and zymogen granules has been observed by measuring the amount of membrane protein, enzymes, and peptides which stay associated with the granules after centrifugation through a 31% sucrose cushion. The interaction was a function of the Ca++ concentration, starting at 1×10−6 m and being saturated at 2×10−5 m of free Ca++ (apparentK m =6.5×10−6 m), and showed preference for Ca++ over other divalent cations with a selectivity sequence (at 0.5mm of total cation concentration): Ca++ 100, Mg++ 35, Ba++ 25, Sr++ 20. The interaction between membranes and granules was specific for cat pancreatic membranes as opposed to cat liver membranes, and for pancreatic zymogen granules as opposed to pancreatic mitochondria. Only 30% of the membrane fraction was bound at saturating levels of zymogen granules and the bound fraction contained alkaline phosphatase, but not other pancreatic plasma membrane markers such as adenylate cyclase or 5′-nucleotidase. After the interaction, removal of Ca++ by the calcium chelator EGTA only partially (about 30%) reversed binding of labelled membranes to the zymogen granules. The process appears to be dependent on the membrane proteins, since brief trypsinization of membranes prior to the assay completely abolished the Ca++-induced interaction. It is concluded that 1) the observed binding may reflect an initial Ca++-dependent event in the process of fusion of zymogen granules with the apical plasma membranes of acinar cells, and 2) protein recognition sites on the interacting membranes are essential for this process.

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Author notes

  1. B. E. Argent

    Present address: Department of Physiology, University Medical School, NE1 7RU, Newcastle upon Tyne, England

  2. G. Sachs

    Present address: Laboratory of Membrane Biology, University of Alabama in Birmingham, 35294, Birmingham, Alabama

Authors and Affiliations

  1. Max-Planck-Institut für Biophysik, 6000, Frankfurt (Main), Germany

    S. Milutinović, B. E. Argent, I. Schulz & G. Sachs

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  1. S. Milutinović
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  3. I. Schulz
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  4. G. Sachs
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Milutinović, S., Argent, B.E., Schulz, I. et al. Studies on isolated subcellular components of cat pancreas. J. Membrain Biol. 36, 281–295 (1977). https://doi.org/10.1007/BF01868155

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  • DOI: https://doi.org/10.1007/BF01868155

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