Lookup NU author(s): Dr Catherine Arden,
Dr Andrew Harbottle,
Professor Loranne Agius
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The association of glucokinase with insulin secretory granules lias been shown by cell microscopy techniques. We used MIN6 insulin-secretory cells and organelle fractionation to determine the effects of glucose on the subcellular distribution of glucokinase. After permeabilization with digitonin, 50% of total glucokinase remained bound intracellularly, while 30% was associated with the 13,000g participate fraction. After density gradient fractionation of the organelles, immunoreactive glucoldnase was distributed approximately equally between dense insulin grannies and low-density organelles that cofractionate with mitochondria. Although MIN6 cells show glucose-responsive insulin secretion, glucokinase association with the granules and low-density organelles was not affected by glucose. Subfractionation of the insulin granule components by hypotonic lysis followed by sucrose gradient centrifugation showed that glucokinase colocalized with the granule membrane marker phogrin and not with insulin. PFK2 (6-phosphofructo-2-kinase-2/fructose-2,6-bisphotase)/FDPase-2, a glucokinase-binding protein, and glyceraldehyde phosphate dehydrogenase, which lias been implicated in granule fusion, also colocalized with glucokinase after hypotonic lysis or detergent extinction of the granules. The results suggest that glucokinase is an integral component of the granule and does not translocate during glucose stimulation.
Author(s): Arden C, Harboottle A, Baltrusch S, Tiedge M, Agius L
Publication type: Article
Publication status: Published
ISSN (print): 0012-1797
ISSN (electronic): 1939-327X
Publisher: American Diabetes Association
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