Purification and characterization of extracellular lipase from a new strain: Pseudomonas aeruginosa SRT 9

Abstract

An extra cellular lipase was isolated and purified from the culture broth of Pseudomonas aeruginosa SRT 9 to apparent homogeneity using ammonium sulfate precipitation followed by chromatographic techniques on phenyl Sepharose CL-4B and Mono Q HR 5/5 column, resulting in a purification factor of 98 fold with specific activity of 12307.8 U/mg. The molecular weight of the purified lipase was estimated by SDS-PAGE to be 29 kDa with isoelectric point of 4.5. Maximum lipase activity was observed in a wide range of temperature and pH values with optimum temperature of 55ºC and pH 6.9. The lipase preferably acted on triacylglycerols of long chain (C14-C16) fatty acids. The lipase was inhibited strongly by EDTA suggesting the enzyme might be metalloprotein. SDS and metal ions such as Hg , Ag 2+ and Fe 2+ decreased the lipase activity remarkedly. Its marked stability and activity in organic solvents suggest that this lipase is highly suitable as a biotechnological tool with a variety of applications including organo synthetic reactions and preparation of enantiomerically pure pharmaceuticals. The Km and Vmax value of the purified enzyme for triolein hydrolysis were calculated to be 1.11 mmol/L and 0.05 mmol/L/min respectively.

Discovery and Characterization of a Thermostable Esterase from an Oil Reservoir Metagenome

“…similar to the observed activity increase of FS02 in the current work, enzymatic activity of an alkaline lipase was shown to be increased in the presence of e.g. methanol [31], and high solvent tolerance of a Pseudomonas lipase has been shown [30]. Hence enzyme activity increase in the presence of solvents observed for FS02 is not unique; however, despite of some uncertainties concerning the absolute activity levels from high standard deviations in the analyses, the increases in activity observed for enzymes in previous studies are rarely as high as observed for FS02, rendering the newly discovered enzyme with distinct features compared to other similar enzymes.…”

“…Lipase activity in the presence of different solvents has been analyzed in several previous studies, mainly showing no or negative effects; e.g. unaffected activity with methanol, ethanol and acetone, and a decreased lipase activity in the presence of isopropanol [30], or reduced lipase activity in the presence of isopropanol, methanol, acetone, acetonitrile and/or ethanol [27] [29]. On the other hand, Figure 2.…”

Applied Biocatalysis with an Organic Resistant Partially Purified Lipase from P. aeruginosa During FAME Production

“…Reported reaction times varied from 8 hours for immobilized enzymes to 90 hours for the same free enzymes, depending on the feedstock and alcohol [25]. Moreover, temperature of 55°C as the working temperature for lipases from P. aeruginosa has been reported [28,29]. This temperature showed, for all the cases, the best balance between stability and reaction rate.…”

“…We carried out a partial purification process that must be improved further according the results. Other researches had reached lipase purification factors of approximately 300, improving their activity approximately 1000-fold compared to the enzyme supernatant [15,28,47]. Also, protein engineering is widely used to improve the lipase production yields and enzymatic activity [48,49].…”

Kinetic modeling, production and characterization of an acidic lipase produced by Enterococcus durans NCIM5427 from fish waste

“…3c). Lipase activity of Pseudomonas aeruginosa MTCC 2488 was drastically reduced in the presence of 2-propanol, hexane and butanol (Borkar et al 2009). Also, the lipase activity of Pseudomonas aeruginosa SRT 9 was drastically reduced in the presence of ethanol, butanol and isopropanol (Chouhan and Dawande 2010).…”