Acyl carrier proteins from developing seeds of Cuphea lanceolata Ait

Abstract

The structure of acyl carrier protein (ACP) may determine the fate of the acyl moieties linked to it in the course of de-novo fatty acid synthesis in higher plants. To investigate a possible correlation between the structure of ACP and the synthesis of medium-chain fatty acids, we isolated and characterized ACP from the seeds of Cuphea lanceolata Ait. (subgenus Eucuphea/Section Heterodon), an annual crop that contains up to 90% decanoic (capric) acid in seed triacylglycerols. After a cell-free extract prepared from developing seeds was treated to 65% saturation with ammonium sulfate, two ACP-isoforms (ACP 1 and ACP 2) were identified in the supernatant that could be purified to homogeneity by anion-exchange chromatography and subsequent reversed-phase high-performance liquid chromatography. The molecular mass determined by matrix-assisted ultraviolet-laser desorption ionization mass spectrometry of ACP 1 was 9315 Da, whereas further heterogeneity was observed for ACP 2 with molecular masses of 8598 and 8703 Da. Aminoterminal sequencing was performed showing a high homology in the primary structures of ACP 1 and ACP 2. Both isoforms were present in the embryo, whereas in the chloroplast-containing seed coat ACP 2 was found in minute amounts, if at all. The expression of ACP 2 correlated with the production of capric acid during the phase of storage-lipid accumulation. These data indicate that ACP 2 is part of the machinery for the synthesis of medium-chain fatty acids, whereas ACP 1 appears to be a constitutive protein.