Abstract
Enzymatic hydration of oleic acid into 10-hydroxystearic acid (10-HSA) represents a theme of substantial scientific and practical interest. In this study, a fatty acid hydratase (OHase) from Lactococcus garvieae was cloned and expressed in Escherichia coli. The recombinantly expressed enzyme was identified as oleate hydratase (EC 4.2.1.53) confirming its highest hydration activity for oleic acid. The optimally yielded enzyme fraction was purified and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). A solitary band on SDS-PAGE confirmed the molecular weight of 65 kDa. Gas chromatography-mass spectrometry (GC-MS) analysis scrutinized the silylated hydroxy fatty acid products acquired from the hydration of oleic acid by the oleate hydratase from L. garvieae. Optimal reaction conditions for the enzymatic production of 10-HSA from oleic acid using the purified oleate hydratase were pH 7.5, 30 °C, 105.49 U/mL enzyme solution and 30 g/L oleic acid. In the presence of activity stimulators, that is, magnesium (II) (Mg2+), the oleate hydratase activity was found to be greatly improved at 30 °C. In conclusion, the results revealed the potential efficacy of recombinant enzyme for the biotechnological conversion of oleic acid to 10-HSA acid with high efficiency. The results would be useful for the improved industrial-scale biosynthesis of 10-HSA via an economical and environmentally friendly bioprocess approach.
Highlights
As a biocatalyst, hydratases have attracted more attention in recent years
The standard of 10-hydroxystearic acid was prepared in ethyl acetate solution at a concentration of
7.5 for min was the most stable fraction at that period revealed that oleate hydratase incubated at pH 7.5 for 30 min was the most stable fraction at that and retained up to 75%
Summary
Hydratases have attracted more attention in recent years. These enzymes catalyze the hydrogenation of unsaturated fatty acids (UFAs) by stereo-selectively adding water to the carbon-carbon double bond [1]. Hydroxyl groups can be incorporated without the requirement for costly cofactors such as recycling or electron releasing groups [2]. These unique characteristics of hydratases render them interesting candidates for numerous applications in industrial bioprocesses. A recent study revealed that oleic acid hydratase necessitates flavin adenosine dinucleotide (FAD) as a cofactor for the production of 10-hydroxystearic acid [2]. Microscopic characterization revealed that the hydratase is an integral membrane protein, and enzymatic
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