Journal Article

Biochemical Characterization of a Marine Pseudoalteromonas citrea-Derived Fatty Acyl-AMP Ligase That Exhibits N-Acyl Amino Acid Synthetic Activity

Wang, Fanghua, Li, Keyan, Deng, Fuli, Wang, Yonghua

2024

Overview

Activation of fatty acids as acyl-adenylates by fatty acid-AMP ligase (FAAL) is a well-established process contributing to the formation of various functional natural products. Enzymatic characterization of FAALs is pivotal for unraveling both the catalytic mechanism and its role in specific biosynthetic pathways. In this study, we recombinantly expressed and characterized a novel FAAL derived from marine Pseudoalteromonas citrea ( Pc FAAL). Pc FAAL was a cold-adapted neutral enzyme, demonstrating optimal activity at 30 °C and pH 7.5. Notably, its specific activity relied on the presence of Mg 2+ ; however, higher concentrations exceeding 10 mM resulted in inhibition of enzyme activity. Various organic solvents, especially water-immiscible organic solvents, demonstrated an activating effect on the activity of Pc FAAL on various fatty acids. The specific activity exhibited a remarkable 50-fold increase under 4% (v/v) n-hexane compared to the aqueous system. Pc FAAL displayed a broad spectrum of fatty acid substrate selectivity, with the highest specific activity for octanoic acid (C8:0), and the catalytic efficiency ( k cat / K m ) for octanoic acid was determined to be 1.8 nM −1 ·min −1 . Furthermore, the enzyme demonstrated biocatalytic promiscuity in producing a class of N-acyl amino acid natural products, as verified by LC-ESI MS. Results indicated that the Pc FAAL exhibits promiscuity towards 10 different kinds of amino acids and further demonstrated their potential value in the biosynthesis of corresponding functional N-acyl amino acids.

• Publisher: Springer US,Springer Nature B.V
• Subject: Amino acids / Amino Acids - chemistry / Amino Acids - metabolism / Bacterial Proteins - chemistry / Bacterial Proteins - genetics / Bacterial Proteins - metabolism / Biochemistry / Biosynthesis / Biotechnology / catalytic activity / Chemistry / Chemistry and Materials Science / class / Enzymatic activity / Enzyme activity / Enzymes / Fatty acids / Fatty Acids - biosynthesis / Fatty Acids - chemistry / Fatty Acids - metabolism / hexane / Hexanes / Hydrogen-Ion Concentration / ligases / Magnesium / n-Hexane / Natural products / Octanoic acid / Organic solvents / Original Article / Pseudoalteromonas / Pseudoalteromonas - enzymology / Pseudoalteromonas - genetics / Pseudoalteromonas citrea / Solvents / Substrate Specificity