Alkyl-dihydroxyacetone phosphate synthase and dihydroxyacetone phosphate acyltransferase form a protein complex in peroxisomes.
Dihydroxyacetone phosphate (GrnP) acyltransferase and alkyl-GrnP synthase are the key enzymes involved in the biosynthesis of ether phospholipids. Both enzymes are located on the inside of the peroxisomal membrane. Here we report evidence for a direct interaction between these enzymes obtained by the use of chemical cross-linking. After cross-linking and immunoblot analysis alkyl-GrnP synthase could be detected in a 210-kDa complex which was located entirely on the lumenal side of the peroxisomal membrane. Two-dimensional SDS/PAGE demonstrated that GrnP-acyltransferase is also cross-linked in a 210-kDa complex. Co-immunoprecipitation confirmed that the two enzymes interact, in a heterotrimeric complex. Furthermore, alkyl-GrnP synthase can form a homotrimeric complex in the absence of GrnP-acyltransferase as was demonstrated by immunoblot analysis after cross-linking experiments with either GrnP-acyltransferase deficient human fibroblast homogenates or recombinant (His)6-tagged alkyl-GrnP synthase. We conclude that alkyl-GrnP synthase interacts selectively with GrnP-acyltransferase in a heterotrimeric complex and in the absence of GrnP-acyltransferase can also form a homotrimeric complex.
Biermann
J
J
Centre for Biomembranes and Lipid Enzymology, Institut for Biomembranes, Utrecht University, The Netherlands.
Just
W W
WW
Wanders
R J
RJ
Van Den Bosch
H
H
D000217:Acyltransferases / Q000172:deficiency / Q000378:metabolism*; D019883:Alkyl and Aryl Transferases / Q000378:metabolism*; D000818:Animals; D005347:Fibroblasts; D006168:Guinea Pigs; D006801:Humans; D015151:Immunoblotting; D008099:Liver / Q000201:enzymology*; D008830:Microbodies / Q000201:enzymology*; D011233:Precipitin Tests; D011487:Protein Conformation; D014357:Trypsin / Q000378:metabolism
D000217:Acyltransferases; C020051:glycerone-phosphate O-acyltransferase; D019883:Alkyl and Aryl Transferases; C019654:alkylglycerone-phosphate synthase; D014357:Trypsin