Measurement of short-chain acyl-CoA dehydrogenase (SCAD) in cultured skin fibroblasts with hexanoyl-CoA as a competitive inhibitor to eliminate the contribution of medium-chain acyl-CoA dehydrogenase.
Short-chain acyl-CoA dehydrogenase (SCAD) deficiency has so far been reported in only very few patients. This is due, in part, to the problems involved in measuring the activity of SCAD unequivocally. The main reason for this difficulty is that butyryl-CoA, the substrate preferably used for SCAD activity measurements, is also dehydrogenated by medium-chain acyl-CoA dehydrogenase (MCAD). Elimination of this contribution can be achieved by means of immune precipitation with a specific MCAD antibody. We now describe a relatively straightforward assay based on the use of gas chromatography/mass spectrometry for detection. The contribution of MCAD to overall butyryl-CoA dehydrogenation was eliminated by adding excess hexanoyl-CoA to the assay medium. The validity of the method developed was checked by SCAD-activity measurements in fibroblasts from an established SCAD-deficient patient.
Niezen-Koning
K E
KE
Department of Pediatrics, University Hospital Groningen, The Netherlands.
Wanders
R J
RJ
Nagel
G T
GT
Sewell
A C
AC
Heymans
H S
HS
D000214:Acyl Coenzyme A / Q000494:pharmacology*; D042964:Acyl-CoA Dehydrogenase; D044944:Acyl-CoA Dehydrogenases / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000172:deficiency*; D001667:Binding, Competitive; D002478:Cells, Cultured; D005347:Fibroblasts / Q000201:enzymology*; D008401:Gas Chromatography-Mass Spectrometry / Q000706:statistics & numerical data; D006801:Humans; D007223:Infant; D012680:Sensitivity and Specificity; D012867:Skin; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; C044181:hexanoyl-coenzyme A; D044944:Acyl-CoA Dehydrogenases; D042964:Acyl-CoA Dehydrogenase