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Conversion of ATP into Other Adenine Nucleotides Within Isolated Islet Secretory Vesicles. Effect of Cyclic AMP on Phosphorus Translocation

Medical Service, Denver Veterans Administration Hospital and Division of Endocrinology, Department of Medicine, University of Colorado Medical School Denver, Colorado 80262

Reprint requests should be sent to Dr. Sussman.

Abstract

The secretory vesicle fraction (24,000 x g) from rat islets was incubated with labelled ATP and radioactivity recovered as adenine nucleotides and inorganic phosphate. The ATP for incubation was labelled in the adenyl moiety with tritium along with ATP labelled with ³²P either in the alpha (ribosyl) or the gamma (terminal) phosphorus position. After a 1-h period of incubation, 51% of the [³H]ATP that underwent conversion was recovered as AMP in the soluble fraction of the secretory vesicle. Forty-three per cent of the radioactivity was recovered as ADP and 5% as cyclic AMP. The insoluble fraction of the secretory vesicle contained substantially less tritium radioactivity- AMP (0.2%), ADP (0.05%), cyclic AMP (0.05%) and ATP (0.20%).

In the insoluble fraction of the secretory vesicle, phosphorus radioactivity was found which could not be accounted for by the presence of adenine nucleotides. The ratio of ³²P to ³H (incubation with ATP labelled with tritium and in the gamma phosphorus position) increased from 0.25 to 0.53 ± 0.041 (P < 0.02). With the addition of cyclic AMP, the ratio increased further to 0.92 ± 0.064 (P < 0.02). The increase of the ³²P to ³H ratio occurred as a consequence of increased recoverable phosphorus in the insoluble fraction.

The secretory vesicle fraction possesses the capability of converting ATP to other adenine nucleotides. As part of the metabolism of ATP, there occurs a translocation of die terminal phosphorus insoluble secretory vesicle constituents. The addition of cyclic AMP (3.5 x 10^–6M) increases phosphorus radioactivity in the insoluble secretory vesicle material to a level above that accounted for by the presence of adenine nucleotides.

Footnotes

Supported by the Medical Research Service of the Veterans Administration Hospital, Denver, Colorado, and the Upjohn Company, Kalamazoo, Michigan.

^* Clinical Investigator, Veterans Administration Hospital, Denver, Colorado (MRIS #8227).

Presented in part at the 57th Annual Meeting of the Endocrine Society, June 18, 1975, New York, New York.

Received January 16, 1976.

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