The activities, properties, and steady-state kinetics of the five
enzymes catalyzing the synthesis of 1-acyl- and 1-alkyl-sn-glycerol 3-phosphate in the cultured skin fibroblasts from
Zellweger syndrome patients and normal controls were studied in detail. Judging from their Km and Vmax values,
glycerol phosphate acyltransferase (EC 2.3.1.15), acyl/alkyl
dihydroxyacetone phosphate reductase (EC 1.1.1.101), and
acyl coenzyme A reductase (long-chain alcohol forming), appear to be affected only slightly by the absence of peroxisomes characteristic of the
Zellweger syndrome.
Glycerophosphate acyltransferase also showed no differences in
N-ethylmaleimide sensitivity nor in inhibition by
dihydroxyacetone phosphate between these cell types.
Dihydroxyacetone phosphate acyltransferase (EC 2.3.1.42) and
alkyl dihydroxyacetone phosphate synthase (EC 2.5.1.26) have altered activity and kinetic constants in homogenates from
Zellweger syndrome fibroblasts.
Dihydroxyacetone phosphate acyltransferase has similar Km (DHAP) values in both control and
Zellweger syndrome cells; however, the value for the Vmax in
Zellweger syndrome cells is only 6% of that found in the controls. This is interpreted as indicating that this
enzyme is not defective in this disease but is simply present at a depressed level. Also, this
enzyme activity has a maximum rate at pH 7.0-7.5 in the mutant cells as opposed to pH 5.4 in the controls. Acylation of
dihydroxyacetone phosphate by control cell homogenate was stimulated by
N-ethylmaleimide at both pH 5.7 and 7.5 whereas this activity from
Zellweger syndrome cells was slightly inhibited at pH 5.7 and strongly inhibited at pH 7.5. In the absence of
detergent,
dihydroxyacetone phosphate acyltransferase in the
Zellweger syndrome cells was much more labile to
trypsin than in the control cells.
Alkyl dihydroxyacetone phosphate synthase had a slightly higher Km (33 vs 17 microM) for
palmitoyl dihydroxyacetone phosphate and a lower Vmax (0.07 vs 0.24 mU/mg
protein) in the
Zellweger syndrome cells as compared to controls. Although this is a substantial decrease in activity, it probably contributes little to the decreased rate of
ether lipid synthesis in these cells. The major problem in this respect is apparently the loss of
dihydroxyacetone phosphate acyltransferase activity. All of these
enzymes, in both control and
Zellweger syndrome cell homogenates, are sedimentable by centrifugation at 100,000g. Also, with the exception of
dihydroxyacetone phosphate acyltransferase they had similar patterns of inactivation by heat in both cell types.