title: A novel halophilic lipase, LipBL, showing high efficiency in the production of eicosapentaenoic acid (EPA).
abstract: BACKGROUND
  Among 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.
  
  
  RESULTS
  A 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.
  
  
  CONCLUSIONS
  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 PUFAs.
authors.affiliation: Department of Microbiology and Parasitology, Faculty of Pharmacy, University of Seville, Seville, Spain.
authors.forename: Dolores
authors.initials: D
authors.lastname: Pérez
authors.forename: Sara
authors.initials: S
authors.lastname: Martín
authors.forename: Gloria
authors.initials: G
authors.lastname: Fernández-Lorente
authors.forename: Marco
authors.initials: M
authors.lastname: Filice
authors.forename: José Manuel
authors.initials: JM
authors.lastname: Guisán
authors.forename: Antonio
authors.initials: A
authors.lastname: Ventosa
authors.forename: María Teresa
authors.initials: MT
authors.lastname: García
authors.forename: Encarnación
authors.initials: E
authors.lastname: Mellado
mesh_terms: D000595:Amino Acid Sequence; D015118:Eicosapentaenoic Acid / Q000096:biosynthesis*; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D004800:Enzymes, Immobilized / Q000378:metabolism; D005395:Fish Oils / Q000378:metabolism; D019766:Genes, Archaeal; D016931:Halobacteriales / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D006868:Hydrolysis / Q000187:drug effects; D008049:Lipase / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008066:Lipolysis / Q000187:drug effects / Q000235:genetics; D008969:Molecular Sequence Data; D000069463:Olive Oil; D010938:Plant Oils / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012965:Sodium Chloride / Q000494:pharmacology; D012997:Solvents / Q000494:pharmacology; D013237:Stereoisomerism; D013379:Substrate Specificity / Q000187:drug effects; D013696:Temperature
chemical_list: D004800:Enzymes, Immobilized; D005395:Fish Oils; D000069463:Olive Oil; D010938:Plant Oils; D012997:Solvents; D012965:Sodium Chloride; D015118:Eicosapentaenoic Acid; D008049:Lipase