Abstract
BackgroundAmong extremophiles, halophiles are defined as microorganisms adapted to live and thrive in diverse extreme saline environments. These extremophilic microorganisms constitute the source of a number of hydrolases with great biotechnological applications. The interest to use extremozymes from halophiles in industrial applications is their resistance to organic solvents and extreme temperatures. Marinobacter lipolyticus SM19 is a moderately halophilic bacterium, isolated previously from a saline habitat in South Spain, showing lipolytic activity.Methods and FindingsA lipolytic enzyme from the halophilic bacterium Marinobacter lipolyticus SM19 was isolated. This enzyme, designated LipBL, was expressed in Escherichia coli. LipBL is a protein of 404 amino acids with a molecular mass of 45.3 kDa and high identity to class C β-lactamases. LipBL was purified and biochemically characterized. The temperature for its maximal activity was 80°C and the pH optimum determined at 25°C was 7.0, showing optimal activity without sodium chloride, while maintaining 20% activity in a wide range of NaCl concentrations. This enzyme exhibited high activity against short-medium length acyl chain substrates, although it also hydrolyzes olive oil and fish oil. The fish oil hydrolysis using LipBL results in an enrichment of free eicosapentaenoic acid (EPA), but not docosahexaenoic acid (DHA), relative to its levels present in fish oil. For improving the stability and to be used in industrial processes LipBL was immobilized in different supports. The immobilized derivatives CNBr-activated Sepharose were highly selective towards the release of EPA versus DHA. The enzyme is also active towards different chiral and prochiral esters. Exposure of LipBL to buffer-solvent mixtures showed that the enzyme had remarkable activity and stability in all organic solvents tested.ConclusionsIn this study we isolated, purified, biochemically characterized and immobilized a lipolytic enzyme from a halophilic bacterium M. lipolyticus, which constitutes an enzyme with excellent properties to be used in the food industry, in the enrichment in omega-3 PUFAs.
Highlights
Lipases (E.C. 3.1.1.1) and carboxylesterases (E.C. 3.1.1.3) constitute the two classes of a/b-fold hydrolases, which are widely distributed in nature from bacteria to higher eucaryotes
In this study we isolated, purified, biochemically characterized and immobilized a lipolytic enzyme from a halophilic bacterium M. lipolyticus, which constitutes an enzyme with excellent properties to be used in the food industry, in the enrichment in omega-3 Polyunsaturated Free Fatty Acids (PUFAs)
To determine the number and the localization of lipolytic enzymes produced by M. lipolyticus SM19, the different cellular fractions of this bacterium were extracted and a zymographic analysis was performed as described in Materials and Methods
Summary
Lipases (E.C. 3.1.1.1) and carboxylesterases (E.C. 3.1.1.3) constitute the two classes of a/b-fold hydrolases, which are widely distributed in nature from bacteria to higher eucaryotes. Since industrial processes are commonly carried out under harsh conditions, it would be of great importance to obtain lipases which retain their optimal activity at extremes of temperature, pH, different concentrations of salts and in the presence of organic compounds normally used in the industrial reactions as solvents [5]. In this sense, the lipases isolated from extreme microorganisms constitute an excellent alternative in the industrial processes [6]. Marinobacter lipolyticus SM19 is a moderately halophilic bacterium, isolated previously from a saline habitat in South Spain, showing lipolytic activity
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