Isolation and characterization of calmodulin from an insulin-secreting tumour.
A major protein constituent of a rat islet cell tumour that exhibited Ca2+-dependent changes in electrophoretic mobility has been purified to homogeneity and compared in its physicochemical and biological properties with bovine brain and rat brain calmodulin (synonymous with phosphodiesterase activator protein, calcium-dependent regulator, troponin C-like protein and modulator protein). The protein, like these calmodulins, contained trimethyl-lysine, exhibited a blocked N-terminus and had an identical amino-acid composition and molecular weight on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Peptide "maps' prepared after digestion of the three proteins with trypsin, papain or Staphylococcus V-8 proteinase were virtually superimposable. Ca2+ altered the electrophoretic mobilities the enhanced the native protein fluorescence in an equivalent manner with all three proteins. Equilibrium dialysis experiments demonstrated in each case the binding of 4g-atoms of calcium/mol of protein; the binding sites were equivalent and showed Kd 0.8 microM. Tumour and brain proteins were equipotent as Ca2+-dependent activators of partially purified rat brain cyclic nucleotide phosphodiesterase, and in this action were inhibited in an identical manner by trifluoperazine. The proteins also exhibited the common property of Ca2+-dependent binding to troponin I, histone H2B and myelin basic protein. The estimated tumour content of calmodulin was 450 mg/kg fresh wt., a value similar to that reported in islets of Langerhans. These results further document the validity of the islet cell tumour as an experimental model of Ca2+-mediated molecular events associated with insulin secretion. They also suggest that brain calmodulin may be substituted for endogenous calmodulin in experimental investigations into the mechanism of insulin secretion.
D007516:Adenoma, Islet Cell / Q000032:analysis*; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001923:Brain Chemistry; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000302:isolation & purification*; D002147:Calmodulin / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D046911:Macromolecular Substances; D010190:Pancreatic Neoplasms / Q000032:analysis*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification; D051381:Rats; D013053:Spectrophotometry
D000596:Amino Acids; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D046911:Macromolecular Substances; D011506:Proteins; D010727:Phosphoric Diester Hydrolases; D002118:Calcium