A 30-year-old woman with clinical features and biochemical findings of muscle phosphofructokinase deficiency was found to have a very low level of alpha-1,4-glucan:alpha-1,4-glucan-6-transglucosylase (branching enzyme, EC 2.4.1.18) activity in muscle. In contrast, branching enzyme activity in the leukocytes was in the range of control values. After sedimentation of the glycogen from muscle homogenates by centrifugation at 105,000 g, branching enzyme activity in muscle of the patient was similar to that of control subjects. This patient illustrates the possibility of falsely diagnosing branching enzyme deficiency when muscle glycogen content is elevated. It is likely that such an artefact may also cause a false positive diagnosis of branching enzyme deficiency in other metabolic diseases associated with glycogen accumulation.

SThe assay of glycogen debranching activity by measurement of the release of glucose from phosphorylase limit dextrin (PLO), W P '. originally used for assay of enzyme activity in muscle and iver tissue. Results are corrected for the presence of nonspecific glucosidases by subtraction of the activity obtained with glycogen as substrate. When assays are performed on muscle or liver, the activity obtained with glycogen is very low and its exact nature is not of practical importance. The widespread use of blood cells and fibroblasts, in which the actiVity on glycogen is considerable, has prompted an examination of the apparent kinetic constants of this reaction. In liver and muscle the apparent Km for PLO ranged from 0.5-2.0 mg/ml, whereas that for glycogen was lower by an order of magnitude. The Vmax with glycogen as substrate did not exceed 20% of that with PLO. In leukocytes and platelets the Km for glycogen was higher than for PLO (0.1 and 0.5 mg/ml for PLO, 0.4 and 0.8 mg/ml for glycogen, in platelets and leukocytes, respectively), and the Vmax for PLO exceeded that for glycogen by 80%. In fibroblasts the Km for both PLO and glycogen was 1.5-3 mg/ml and the differences in Vmax were small. These results indicate that substrate concentration should be varied according to the kinetic constants of each cell type and point to the importance of distingUishing between the (low?) activity of the debranching enzyme on glycogen and nonspecific hydrolysis. tally by stable isotope dilution GCMS assays; there are 15 others potentially diagnosable. This methodology has been applied to mevalonic aciduria, the first documented inherited disorder of cholesterol and nausterol isoprene biosynthesis in man. In a mother at risk for this disease the urinary excretion of mevalonic acid at 16 weeks of pregnancy of 5.6 mmol/mol creatinine was 35 imes the mean normal level (range 0.09 -0.22 mmol/mol creatinine, 5 control females). The analysis of amniotic fluid at this time indicated a 3000-fold elevation of mevalonic acid (240 pmolA; range in 4 control amniotic fluids: 0.054 -0.11 pmol/l), indicating the presence of an affected fetus. The diagnosis was confirmed by demonstration of deficient mevalonate kinase 'in amniocytes and ultimately in liver from the abortus. Mevalonic acid was found to be highly elevated in the abortus tissues. Concentrations ranged from 840 to 1120 pmol/kg in adrenals, gonads, liver, lymph nodes and spleen. An even higher level was detected in the brain, where the concentration was 1810 ,mol/kg (control1 pmol/kg). This may reflect a particular need for cholesterol in brain development.and is consistent with the severe developmental delay of the index patient with mevalonic aciduria. After termination of the pregnancy the mother's urinary excretion of mevalonic acid fell to 0.20 mmol/mol creatinine, within the normal range. I 64ASSAY OF AMYLO-I,6-GLUCOSIDASE-TRANSFERASE ACTIVITY Mevalonate aciduria is an inherited disorder of cholesterol synthesis. Mevalonate kinase activity in lysates of fibroblasts fr...