Purification and characterization of thermostable and alcohol tolerant lipase from Pseudoxanthomonas sp.

Abstract

An extracellular lipase from Psedoxanthomonas sp. was purified 47.3 folds with an overall yield of 27% through purification procedure of acetone precipitation, ion exchange and gel filtration chromatography. Protein precipitation using acetone fractionation showed that the enzyme was precipitated at the fraction of 0 to 40%. Further purification of the enzyme by ion exchange followed by gel filtration chromatography showed that there were two types of lipases with similar molecular size at around 50 kDa. Characterization of optimum pH, temperature, and substrate specificity were carried out against the isolated lipase. Lip1 showed specific substrate preferences towards p-nitro phenyl myristate meanwhile Lip2 exhibited hydrolytic activity towards short and medium acyl chain of the substrate. Further analysis of both enzyme activities on variation of pH and temperature showed that the optimum pH and temperature for Lip1 were at pH 10.0 and 70°C, respectively while pH 8.0 and 50°C were the optimum pH and temperature for Lip2 respectively. Lyophilized lipase isolated from acetone fraction showed lipolytic activity under variation of methanol concentration up to 30%.

Effect of mutation at oxyanion hole residu (H110F) on activity of Lk4 lipase

“…Our previous report showed that lipase from Pseudoxantomonas sp. still exhibited an activity in the presence of methanol up to 30 % concentration [ 37 ]. Furthermore, lipase from Geobacillus thermoleovorans showed tolerance to various polar solvents and loss of activity in the presence of n-hexane and chloroform [ 38 ].…”

Thermostable Lipase from Domestic Compost Isolated Bacteria

“…The result showed that the maximum activity at pH 9 ( Fig. 3), suggested that the enzymewas an alkali tolerant lipase 10,13 . The high activity of the isolated at alkaline condition suggests that the lipase shows high potential application on many industries such as detergent, biopolymers, pharmacy, cosmetics and flavor 17 .…”

“…Isolate AL89 was inoculated in 100 mL media (0.1% CaCl 2 , 0.5% yeast extract, 0.5% meat extract, 0.1% NaCl in 0.05 M phosphate buffer pH 8.0),then incubated at 55 o C 150 rpm for 19 hours, as was done in a previous study 13 . Extracellular lipase was obtained from culture supernatants which had been centrifuged for 45 minutes at 12000 g.…”

“…The lipolytic activity assay for this lipase was carried out by the spectrophotometric method as explained by Syihab (2017) 13 .…”

“…Lipase from Thermus aquaticus, isolated from Indonesian hot spring, was obtained through homologous expression and purified by hydrophobic interactions-based chromatography 12 . Thermostable lipases from Pseudoxanthomonas sp., isolated from domestic compost was reported and showed tolerance towards methanol and various organic solvent 13 .…”