Work

Biochemical and structural analysis of -AtN-Myc down regulated like-1 reveals phosphatidic acid regulated esterase function

Year: 2026

Type: article

Abstract: The N-Myc Downregulated-Like (NDL) protein family in Arabidopsis was identified as an interacting partner of components of the G-protein complex. NDL's role has been established in regulating auxin transport through modulation of auxin transport facilitators in an Arabidopsis G-protein β subunit (AGB1)-dependent manner. NDL proteins are modulators of abiotic stress response, such as high salt and drought, and may serve as predictors of plant morphophysiological responses based on their gene expression patterns. The domain organization of NDL proteins predicts that they contain esterases/lipases with an α/β hydrolase fold. Little is known of this protein family. Biochemical analysis of Arabidopsis NDL proteins was conducted to understand the mechanism underlying their role in G protein-mediated abiotic stress-related functions and to guide hypothesis-driven experiments for long-term crop genetic improvement. To determine whether NDL1 is a lipase/esterase, we first optimized the expression of recombinant NDL1 in E. coli and tested it structurally and biochemically for lipase/esterase activity. We further analyzed NDL1’s substrate-binding preference using different lipids to elucidate its mode of action in abiotic stress. We demonstrated that purified NDL1 exhibits phosphatidic acid (PA)-competitive hydrolysis of short-chain substrates, such as p-nitrophenyl acetate and p-Nitrophenyl butyrate, but not of long-chain substrates, p -Nitrophenyl laurate and p -Nitrophenyl palmitate, confirming esterase activity. We identified strong binding of NDL1 to the membrane PA and ceramide-1-phosphate, as well as weak binding towards five anionic glycerophospholipids. PA's binding and inhibition of esterase activity suggest that lipid-mediated regulation of NDL1 function is involved. • Arabidopsis N-Myc Downregulated-Like proteins are interactors of Gβγ and RGS1 • NDL proteins are possible abiotic stress predictive markers • NDL1 belongs to the lipase/esterase superfamily containing an α/β hydrolase fold • Lipid Overlay shows strong binding to Phosphatidic acid and Ceramide-1-phosphate • N-terminal modified NDL1 exhibits Esterase activity • NDL1 shows PA-competitive inhibition of its Esterase activity • Docking of NDL1 with PNPA, PA and C1P predicted four shared amino acids

Source: Plant Physiology and Biochemistry

DOI: 10.1016/j.plaphy.2026.111380