STUDY OF Ca2+ ION EFFECT ON LIPASE ACTIVITY OF HALOPHILIC BACTERIA Chromohalobacter japonicus BK-AB18

• Main Author: Widhyastuti, Henny
• Format: Theses
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/34036
• Institution: Institut Teknologi Bandung

Lipase (triacylglycerol acylhydrolases, EC 3.1.1.3), which is classified as lipolytic enzymes is commonly applied in variety of industrial biotechnology. As biocatalyst, lipase play role in catalizing hydrolysis of triacyl glycerol and esterification of fatty acids. Screening of lipase-producing microbes have been used as one of the way to obtain novel varians of lipases that may stable at high temperatures, pH, high salinity and organic solvents. Halophilic microbes that can live in hypersaline environment has been known to be a potential source to produce enzymes that are not only stable against high concentration of salt, but also have other uniques properties such as stable in various types of solvent ranging from polar to nonpolar ones. In previous studies, local halophilic bacteria have been isolated from "Bledug Kuwu" mud crater that periodically spitting brine to the surface creating hypersaline environment. In this study, halophilic bacteria, Halomonas and Chromohalobacter, isolated from this habitate were selected to be explored its potential in producing extracellular lipase. The screening results in various NaCl concentrations found that Chromohalobacter japonicus BK-AB18 was the most potential isolate among the others in producing extracelular lipase. Zymography and hydrolytic activity assay suggested that extracellular lipase was found at 60-70% of ammonium sulfate fraction. The molecular weight of this lipase was about 37 kDa. This enzyme exhibited optimum activity at 45 °C and pH 9.5. The addition of various metalic ions affecting lipase activity and among those, Ca2+ ion was the most significant in increasing the enzyme activity. Lipase activity decreased significantly in the presence of PMSF (serine hydrolase inhibitor) and SDS (detergent). This enzyme also exhibited wide tolerance on various types of organic solvent. Its activity increased significantly in n-propanol, n-butanol, isopropanol, acetonitrile and chloroform. Addition of Ca2+ ion to the enzyme solutions enhanced its activity particularly in n-hexane, SDS, and 8% NaCl solution. In additions, the presence of Ca2+ ion also stabilized the enzyme when incubated for 9 hours at 45 °C and 55 °C. Overall, stability and activity of lipase isolated from halophilic bacteria Chromohalobacter japonicus BK-AB18 was enhanced by the presence of Ca2+ ion