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A cold tolerant lipase develops enhanced activity, thermal tolerance and solvent stability in the presence of calcium nanoparticles: An alternative approach to genetic modulation

Das, Aniket, Chakrabarti, Krishanu
Biocatalysis and agricultural biotechnology 2018 v.15 pp. 1-10
Pseudomonas aeruginosa, bacteria, barium, butyrates, calcium, calcium chloride, carboxylic ester hydrolases, cobalt, cold, cold tolerance, enzyme activity, esterification, fatty acid esters, half life, heat, heat tolerance, hexane, iron, long chain fatty acids, magnesium, molecular weight, nanoparticles, nickel, pH, short chain fatty acids, soil, solvents, sunflower oil, temperature, zinc
An extracellular cold active lipase producing bacterium was isolated from soil. It was identified as Pseudomonas aeruginosa KC1 strain (GenBank accession number KT334371). There are no previous reports on purification of cold active lipase protein from Pseudomonas aeruginosa. The lipase of molecular weight 54 kDa was purified to 33 fold with 8% recovery. The enzyme was active within the range 10–40 °C with maximum activity at 15 °C; pH 7.0–8.5 with 4-nitrophenyl butyrate substrate. Substrate utilization by lipase showed better affinity for short chain fatty acid esters. The enzyme activity was enhanced by Ca2+, Ba2+, Fe2+ and Mg2+; inhibited by Hg2 +, Ni2+, Zn2+ and Co2+. Nano-calcium enabled KC1 lipase (NP-lip) showed enhanced activity for both short and long chain fatty acid esters (NP: 8.8 µg/mL) compared to CaCl2 (1 mM). The activity of the NP-lip system increased 72% at 15 °C and 7 fold at 55 °C while retaining activity for 4 h. Lowering of Km (55% at 15 °C; 45% at 55 °C) and increased Vmax (7 fold at 15 °C; 3.5 fold at 55 °C) was observed for NP-lip system. Heat deactivation kinetics for NP-lip showed drastic improvement in half-life at higher temperatures and entropy-enthalpy compensation. Furthermore the NP-lip system was stable in organic solvents and was effective in the esterification of butyl butyrate and trans-esterification of sunflower oil in n-hexane. This remarkable simultaneous enhancement of activity, temperature and organic solvent tolerance of the NP-lip has potential for industrial usage.