Induction of Ca2+ transport in liposomes by insulin.
The requirement of extracellular Ca2+ for insulin action has been indicated by past studies. With a view to understand the interaction of insulin with Ca2+ in the vicinity of the cell membrane, we have examined the ability of insulin and its constituent polypeptide chains A and B to translocate Ca2+ and Mg2+ across the lipid bilayer in two sets of synthetic liposomes. The first were unilamellar vesicles made of dimyristoylphosphatidylcholine and contained the Ca2+ sensor dye arsenazo III. Peptide-mediated Ca2+ and Mg2+ transport in these vesicles was monitored at 37 degrees C in a neutral buffer containing CaCl2 or MgCl2 using a difference absorbance method. In the second set, multilamellar vesicles of egg lecithin containing trapped fura-2 were employed and the cation transport was followed at 20 degrees C by fluorescence changes in the dye. Control experiments indicated that the hormonal peptides caused no appreciable perturbation of the vesicles leading to leakage of contents or membrane fusion. In both liposome systems, substantial Ca2+ and Mg2+ transport was observed with insulin and the B chain; the A chain was less effective as an ionophore. Quantitative analysis of the transport kinetic data on the B chain showed a 1:1 peptide-Ca2+ complex formed inside the membrane. In light of the available structural data on Ca2+ binding by insulin and insulin receptor, our results suggest the possibility of the hormone interacting with the receptor with the bound Ca2+.
Brimble
K S
KS
Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada.
Ananthanarayanan
V S
VS
D001692:Biological Transport; D002118:Calcium / Q000378:metabolism*; D004134:Dimyristoylphosphatidylcholine / Q000378:metabolism; D007328:Insulin / Q000502:physiology*; D007700:Kinetics; D008081:Liposomes / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism
D007328:Insulin; D008081:Liposomes; D008274:Magnesium; D002118:Calcium; D004134:Dimyristoylphosphatidylcholine