Characterization of a Novel Acyl-ACP Δ Desaturase Gene Responsible for Palmitoleic Acid Accumulation in a Diatom

Palmitoleic acid (16:1Δ) possesses a double bond at the seventh carbon atom from methyl end of the acyl chain and belongs to unusual ω-7 monounsaturated fatty acids with broad applications in food, pharmaceuticals, cosmetics, biofuel, and other industries. This high-value fatty acid accumulates up to >40% of total lipid in the marine diatom The present study was conducted to determine the key gene responsible for 16:1Δ biosynthesis in this unicellular alga. A new full-length cDNA and genomic DNA encoding acyl-ACP Δ desaturase (PtAAD) were isolated from cells. Expression levels of gene under normal and stress culture conditions were both positively correlated with 16:1Δ accumulation, implying its potential role for fatty acid determination. Functional complementation assay of a yeast mutant strain evidenced that could restore the synthesis of unsaturated fatty acid, especially generating high levels of 16:1Δ. Further transient expression of gene in leaves was accompanied by the accumulation of 16:1Δ, which was absent from control groups. Three-dimensional structure modeling studies showed that functional domain of PtAAD contained three variant amino acids (F160, A223, and L156), which may narrow the space shape of substrate-binding cavity to ensure the entry of 16:0-ACP. Consistent with this prediction, the mutated version of gene (F160L, A223T, and L156M) in systems failed to accumulate 16:1Δ, but increased levels of 18:1Δ. Taken together, PtAAD exhibits a strong enzymatic activity and substrate preference for 16:0-ACP, acting as the key player for high biosynthesis and accumulation of 16:1Δ in this alga. These findings provide new insights for better understanding the palmitoleic acid and oil biosynthetic mechanism in , indicating that gene may have practical applications for enriching palmitoleic acid and oil yield in other commercial oleaginous algae and crops.