Heterogeneity of defects in mitochondrial acetoacetyl-CoA thiolase biosynthesis in fibroblasts from four patients with 3-ketothiolase deficiency.
Deficient mitochondrial acetoacetyl-CoA thiolase in fibroblasts from four patients with 3-ketothiolase deficiency was studied using immunochemical methods. We also examined fibroblasts from two heterozygotes, the mother and the brother of the case 1 patient, identified on the basis of the results of the enzyme activity measurements, using 2-methylacetoacetyl-CoA as substrate. The results were as follows: 1) in fibroblasts from all four patients, the thiolase activity using acetoacetyl-CoA was not activated by K+, although that of the controls and the heterozygotes was activated about 2-fold. 2) by immunoblot analyses, mitochondrial acetoacetyl-CoA thiolase was not detectable in fibroblasts from cases 2 and 3, although a very faint band was seen in tissues from cases 1 and 4. However, the band of mitochondrial 3-ketoacyl-CoA thiolase was clearly detected in all patients to the same extent as in the controls. 3) mitochondrial acetoacetyl-CoA thiolase was observed after pulse labeling for 1-h and a 72-h chase period in three cell lines (cases 1, 2, and 4), but was fainter compared to the controls. In another cell line (case 3), a fluorographic band at the same position was detected following a 1-h pulse, but disappeared following a 6-h chase. These results demonstrate heterogeneity in the enzyme defect resulting in a deficiency of mitochondrial acetoacetyl-CoA thiolase in fibroblasts from patients with 3-ketothiolase deficiency.
Nagasawa
H
H
Department of Pediatrics, Gifu University School of Medicine, Japan.
Yamaguchi
S
S
Orii
T
T
Schutgens
R B
RB
Sweetman
L
L
Hashimoto
T
T
D000101:Acetyl-CoA C-Acetyltransferase / Q000032:analysis / Q000096:biosynthesis*; D000102:Acetyl-CoA C-Acyltransferase / Q000032:analysis / Q000172:deficiency*; D000123:Acetyltransferases / Q000096:biosynthesis*; D000217:Acyltransferases / Q000172:deficiency*; D000328:Adult; D002460:Cell Line; D002648:Child; D005260:Female; D005347:Fibroblasts / Q000201:enzymology / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007223:Infant; D008297:Male; D008928:Mitochondria / Q000201:enzymology* / Q000378:metabolism
D000217:Acyltransferases; D000123:Acetyltransferases; D000102:Acetyl-CoA C-Acyltransferase; D000101:Acetyl-CoA C-Acetyltransferase