Effect Of Zwitterionic Compound On Activity And Stability Of Lipase

In this paper, effect of sulfobetaine methacrylate(SBMA) on activity, stability and conformation of lipase was investigated. The purpose of the research was to explore the role of zwitterionic compound on enzyme.SBMA at different concentrations was added to lipase solution. Very low(0.1mM) or high concentration(1M) of SBMA could increase the lipase activities by25.0%and99.2%respectively. Stability of lipase improved significantly. After96h in37℃, lipase solution became turbid and its relative activity was0.310, but lipase solution with1M SBMA remained clear and its relative activity was1.350. After1h incubated in60℃, the relative activity of lipase was0.154, but the relative activity of lipase with1M SBMA was1.133. These results supported that zwitterion maintained natural enzyme conformation.Amino groups of lipase were modified by a mild vinylation with N-acryloxysuccinimide (NAS), then SBMA monomer was added into the vinylated lipase. The redox polymerization was initiated by free radicals and the modified lipase was produced. The optimumtemperature of modified lipase was55℃. In55℃, the relative activity of modified lipase was1.397, but the relative activity of lipase was0.276. In the condition of pH=3, pH=7.5and pH=10.9. the relative activity of lipase was0.542,1.000and0.433respectively, and the relative activity of modified lipase was0.838,1.397and0.633. UV spectrum of lipase and modified lipase showed that conformation of lipase changed in the condition of high temperature, acid and alkaline. Fluorescence spectra showed that modification did not change natural conformation of lipase.Preparation of encapsulated lipase:SBMA monomer and N,N’-methylene diacrylamide crosslinker were added into vinylated lipase. Then the redox polymerization was initiated by free radicals and the encapsulated lipase was produced.The modification ratio of amino groups of lipase was94.5%. When mass of SBMA monomer was6g and mass ratio between crosslinker and SBMA was7%, encapsulated lipase obtained the best activity yield. In n-heptane, the optimum temperature of encapsulated lipase was40℃. Suitable water content could improve activity of lipase. When water content was2%(V/V), the maximum substrate conversion rate of encapsulated lipase reached93.7%in24h. The activity of encapsulated lipase in6h reached up to2.29-fold of that of lipase. Increasing the substrate concentration could prolong the reaction time and descrease the equilibrium conversion rate. When concentration of lauric acid increased from40mM to200mM, the maximum conversion rate of encapsulated lipase decreased from93.1%to86.4%, but the maximum conversion rate of lipase decreased from91.6%to53.6%. It showed that increasing the concentration of the substrate was better for the encapsulated lipase than lipase. The encapsulated lipase retained77.9%of its initial activity after six recycles.