Purification, Characterization and Immobilization of Lipase from Proteus vulgaris OR34 for Synthesis of Methyl Oleate

Abstract

A newly isolated strain, Proteus vulgaris OR34, from olive mill waste was found to secrete an alkaline extracellular lipase at 11 U• ml -1 when cultivated on an optimized liquid medium. This lipase was purified 94.64fold with a total yield of 9.11% and its maximal specific activity was shown to be 3232.58 and 1777.92 U•mg -1 when evaluated using the pH-stat technique at 55℃ and pH 9 and Tributyrin TC4 or olive oil as the substrate. The molecular mass of the pure OR34 lipase was estimated to be around 31 kDa, as revealed by SDS-PAGE and its substrate specificity was investigated using a variety of triglycerides. This assay revealed that OR34 lipase preferred short and medium chain fatty acids. In addition, this lipase was stable in the presence of high concentrations of bile salt (NaDC) and calcium ions appear not to be necessary for its activity. This lipase was inhibited by THL (Orlistat) which confirmed its identity as a serine enzyme. In addition, the immobilization of OR34 lipase by adsorption onto calcium carbonate increased its stability at higher temperatures and within a larger pH range. The immobilized lipase exhibited a high tolerance to organic solvents and retained 60% of its activity after 10 months of storage at 4℃. Finally, the OR34 lipase was applied in biodiesel synthesis via oleic acid mediated esterification of methanol when using hexane as solvent. The best conversion yield (67%) was obtained at 12 h and 40℃ using the immobilized enzyme and this enzyme could be reused for six cycles with the same efficiency.

Progress and perspectives of enzymatic preparation of human milk fat substitutes

“…However, Novozym 435 exhibited preference for short chain fatty acids and long chain fatty acids, as reported by Wu et al [62]. Duan et al [63], Misbah et al [64] and Rivero-Pino et al [65] found that many lipases had substrate specificity toward triacylglycerols with short or medium carbon chain during the hydrolysis of triacylglycerols. While for the hydrolysis of fatty acid esters, lipases expressed different selectivity.…”

Effect of Pomegranate Peel Extract on the Production of Some Enzymes in Proteus spp Isolated from Different Clinical Samples in Kirkuk city.

“…Protease enzymes are one of the virulence factors for proteus [9]. One of these types of protein-degrading enzymes is the Metalloprotease enzyme, where this enzyme works to analyze proteins that contain metal ions in their composition, such as Na+, K+, which are essential in pathogenesis [10] Proteus secretes the enzyme lipase [11] [12], which includes lipolytic enzyme (general) Lipases and specialized enzyme phospholipases. Lipase enzymes work to break down the ester bonds that bind fatty acids with glycerol, producing a molecule of glycerol and fatty acids [3].…”