Molecular studies of liver aldolase B in hereditary fructose intolerance using blotting and immunological techniques.
Hereditary fructose intolerance is due to a deficiency of liver aldolase (aldolase B). Little is known about its molecular mechanisms. We have tried to demonstrate the presence of the molecule and have explored the possibility of genetic heterogeneity. Liver samples from fifteen cases of hereditary fructose intolerance due to aldolase B deficiency were studied by various electrophoretic techniques. After electrophoresis on polyacrylamide gels, proteins were electrophoretically transferred on to nitrocellulose filters. They were treated with specific antialdolase B antibodies, and then with radioiodinated protein A, followed by autoradiography. Investigations included: (a) sodium dodecyl sulphate electrophoresis, in order to detect the presence of immunologically reactive molecules and to estimate the subunit size; (b) attempts to discover charge anomalies of the native molecule and of its subunits, by the use of: Isoelectric focusing of the native enzyme. Isoelectric focusing and non-equilibrium pH gradient electrophoresis (NEPHGE) after dissociation in urea. The major results were the following: (1) In all cases a cross-reacting material was found, with a molecular subunit size of 38000, indistinguishable from that of controls. (2) Evidence for molecular heterogeneity of the disease was provided by two types of data: amount of apparent immunologically reactive protein, which varied from less than 3% to 100% of that of controls; and charge data, aldolase B from seven patients showing an increased negative charge and from one patient a normal charge.
D002239:Carbohydrate Metabolism, Inborn Errors / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005633:Fructose Intolerance / Q000201:enzymology / Q000235:genetics*; D005634:Fructose-Bisphosphate Aldolase / Q000032:analysis*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007158:Immunologic Techniques; D007223:Infant; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000032:analysis*; D008099:Liver / Q000201:enzymology*; D008297:Male; D008970:Molecular Weight
D007527:Isoenzymes; D005634:Fructose-Bisphosphate Aldolase