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Acyl carrier proteins from sunflower (Helianthus annuus L.) seeds and their influence on FatA and FatB acyl-ACP thioesterase activities

Abstract

Main conclusion

The kinetics of acyl-ACP thioesterases from sunflower importantly changed when endogenous ACPs were used. Sunflower FatB was much more specific towards saturated acyl-ACPs when assayed with them.

Acyl carrier proteins (ACPs) are small (~9 kDa), soluble, acidic proteins involved in fatty acid synthesis in plants and bacteria. ACPs bind to fatty acids through a thioester bond, generating the acyl-ACP lipoproteins that are substrates for fatty acid synthase (FAS) complexes, and that are required for fatty acid chain elongation, acting as important intermediates in de novo fatty acid synthesis in plants. Plants, usually express several ACP isoforms with distinct functionalities. We report here the cloning of three ACPs from developing sunflower seeds: HaACP1, HaACP2, and HaACP3. These proteins were plastidial ACPs expressed strongly in seeds, and as such they are probably involved in the synthesis of sunflower oil. The recombinant sunflower ACPs were expressed in bacteria but they were lethal to the prokaryote host. Thus, they were finally produced using the GST gene fusion system, which allowed the apo-enzyme to be produced and later activated to the holo form. Radiolabelled acyl-ACPs from the newly cloned holo-ACP forms were also synthesized and used to characterize the activity of recombinant sunflower FatA and FatB thioesterases, important enzymes in plant fatty acids synthesis. The activity of these enzymes changed significantly when the endogenous ACPs were used. Thus, FatA importantly increased its activity levels, whereas FatB displayed a different specificity profile, with much high activity levels towards saturated acyl-CoA derivatives. All these data pointed to an important influence of the ACP moieties on the activity of enzymes involved in lipid synthesis.

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Abbreviations

ACP:

Acyl carrier protein

DAF:

Days after flowering

DAG:

Diacylglycerol

FAS:

Fatty acid synthases

PBS:

Phosphate-buffered saline

TAG:

Triacylglycerol

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Acknowledgments

This work was funded by the “Ministerio de Economia y Competitividad” and FEDER (Project AGL2014-53537-R and JAE-CSIC to J.A. A-M).

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Correspondence to Joaquín J. Salas.

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Aznar-Moreno, J.A., Venegas-Calerón, M., Martínez-Force, E. et al. Acyl carrier proteins from sunflower (Helianthus annuus L.) seeds and their influence on FatA and FatB acyl-ACP thioesterase activities. Planta 244, 479–490 (2016). https://doi.org/10.1007/s00425-016-2521-7

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Keywords

  • Acyl carrier protein
  • Sunflower, Helianthus annuus
  • Acyl-ACP thioesterase
  • Fatty acid synthesis