Regioselectivity mechanism of the Thunbergia alata Δ6-16:0-acyl carrier protein desaturase

• DOI: 10.1093/plphys/kiab577
• Publication Date: 2021-12-07T20:18:07Z
• Authors: Jodie E Guy; Yuanheng Cai; Marcel D Baer; Edward Whittle; Jin Chai; Xiao-Hong Yu; Ylva Lindqvist; Simone Raugei; John Shanklin
• Keywords: Acyl carrier protein; Acyltransferases

Abstract Plant plastidial acyl–acyl carrier protein (ACP) desaturases are a soluble class of diiron-containing enzymes that distinct from the integral membrane found in plants and other organisms. The archetype this is stearoyl-ACP desaturase which converts into oleoyl (18:1Δ9cis)-ACP. Several variants expressing regioselectivity have been described including Δ6-16:0-ACP black-eyed Susan vine (Thunbergia alata). We solved crystal structure T. alata at 2.05 Å resolution. Using molecular dynamics (MD) simulations, we identified low-energy complex between 16:0-ACP would position C6 C7 acyl chain adjacent to diiron active site. model was used identify mutant could convert Δ6 Δ9 regioselectivity. Additional modeling ACP confirmed predicted To validate in-silico predictions, synthesized two analyzed their reaction products using gas chromatography-coupled mass spectrometry. Assay results mutants designed selectivity exhibited changes. In complementary experiments, castor lost some desaturation ability gained desaturate position. computational workflow for revealing mechanistic understanding presented herein lays foundation designing acyl-ACP with novel selectivities increase diversity monoenes available bioproduct applications.