%0 Journal Article %A Batumalaie, Kalaivani %A Khalili, Elham %A Mahat, Naji Arafat %A Huyop, Fahrul Zaman %A Abdul Wahab, Roswanira %D 2017 %T A statistical approach for optimizing the protocol for overexpressing lipase KV1 in Escherichia coli: purification and characterization %U https://tandf.figshare.com/articles/journal_contribution/A_statistical_approach_for_optimizing_the_protocol_for_overexpressing_lipase_KV1_in_i_Escherichia_coli_i_purification_and_characterization/5632009 %R 10.6084/m9.figshare.5632009 %2 https://tandf.figshare.com/ndownloader/files/9808450 %K Acinetobacter haemolyticus %K central composite design %K lipase %K protein overexpression %K response surface methodology %X

Lipase is one of the most important industrial enzymes, widely used in the preparation of food additives, cosmetics and pharmaceuticals. In order to obtain a large amount of lipase, in the present study, a gene encoding intracellular lipase was cloned from Acinetobacter haemolyticus. The recombinant lipase KV1 containing a His-tag was expressed in Esherichia coli BL21 (DE3) cells, using pET-30a as the expression vector. Using the central composite design, screening and optimization of induction conditions (cell density before induction, IPTG (isopropyl β-D-1-thiogalactopyranoside) concentration, post-induction temperature and post-induction time) were made. All parameters significantly (P < 0.05) influenced the expression of lipase KV1, rendering a 70% increase in enzyme production at optimum induction conditions (OD600 before induction: 0.6, IPTG concentration: 0.5 mmol/L, post-induction temperature: 40 °C, post-induction time: 16 h). The expressed recombinant lipase KV1 was purified using Ni-affinity chromatography, affording ∼3.1-fold of the enzyme with an estimated relative molecular mass of 39 kDa. The recombinant lipase KV1 exhibited its maximum activity at 40 °C and pH 8.0. Beneficially, the recombinant lipase KV1 retained its relative activities (>80%) even up to 24 h between pH 7−12; suggesting that the recombinant lipase KV1 may be suitable for a wide range of industrial applications.

%I Taylor & Francis