Functional and structural characterization of a novel plastidial FatB2 Acyl-ACP thioesterase from sunflower...

FatB acyl-ACP thioesterases play an important role in plant fatty acid metabolism, determining the chain length and saturation of acyl groups that feed both storage oil and membrane lipid biosynthesis. In sunflower (Helianthus annuus L.), only one isoform, HaFatB1, has been characterized to date. Here, we report the identification and functional characterization of a novel sunflower plastidial thioesterase, HaFatB2, which exhibits low expression in developing seeds but is relatively abundant in leaves, suggesting a metabolic role beyond oil biosynthesis. Structural modelling and molecular docking predicted efficient accommodation of palmitoyl- and oleoyl-ACPs, a preference confirmed by in vitro assays and kinetic studies. When expressed in Escherichia coli, HaFatB2 markedly modified the fatty acid profile, leading to the unexpected accumulation of 4-hydroxymyristic acid (23% of total FAs), a rare fatty acid with potential industrial relevance. In contrast, expression in the photosynthetic cyanobacterium Anabaena sp. PCC 7120 and in Nicotiana benthamiana leaves did not significantly alter fatty acid composition, likely due to differences in substrate availability and endogenous regulatory mechanisms. Altogether, these findings identify HaFatB2 as a thioesterase with high preference for the export of palmitic and oleic fatty acids and highlight its biotechnological potential for producing uncommon hydroxylated fatty acids in heterologous systems.