ABSTRACT
Lipases are enzymes of immense industrial relevance, and, therefore, are being intensely investigated. In an attempt to characterize lipases at molecular level from novel sources, a lipase gene from Bacillus amyloliquefaciens PS35 was cloned, heterologously expressed in Escherichia coli DH5α cells and sequenced. It showed up to 98% homology with other lipase sequences in the NCBI database. The recombinant enzyme was then purified from E. coli culture, resulting in a 19.41-fold purification with 9.7% yield. It displayed a preference for long-chain para-nitrophenyl esters, a characteristic that is typical of true lipases. Its optimum pH and temperature were determined to be 8.0 and 40 °C, respectively. The half-lives were 2.0, 1.0 and 0.5 h at 50 °C, 60 °C and 70 °C, respectively. The metal ions K+ and Fe3+ enhanced the enzyme activity. The enzyme displayed substantial residual activity in the presence of various tested chemical modifiers, and interestingly, the organic solvents, such as n-hexane and toluene, also favored the enzyme activity. Thus, this study involves characterization of B. amyloliquefaciens lipase at molecular level. The key outcomes are novelty of the bacterial source and purification of the enzyme with desirable properties for industrial applications.
Subject(s)
Bacillus/enzymology , Bacillus/genetics , Cloning, Molecular , Gene Expression , Lipase/genetics , Lipase/metabolism , Recombinant Proteins/metabolism , Enzyme Stability , Escherichia coli/genetics , Hydrogen-Ion Concentration , Lipase/chemistry , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Sequence Analysis, DNA , Sequence Homology , Substrate Specificity , TemperatureABSTRACT
Abstract Lipases are enzymes of immense industrial relevance, and, therefore, are being intensely investigated. In an attempt to characterize lipases at molecular level from novel sources, a lipase gene from Bacillus amyloliquefaciens PS35 was cloned, heterologously expressed in Escherichia coli DH5α cells and sequenced. It showed up to 98% homology with other lipase sequences in the NCBI database. The recombinant enzyme was then purified from E. coli culture, resulting in a 19.41-fold purification with 9.7% yield. It displayed a preference for long-chain para-nitrophenyl esters, a characteristic that is typical of true lipases. Its optimum pH and temperature were determined to be 8.0 and 40 °C, respectively. The half-lives were 2.0, 1.0 and 0.5 h at 50 °C, 60 °C and 70 °C, respectively. The metal ions K+ and Fe3+ enhanced the enzyme activity. The enzyme displayed substantial residual activity in the presence of various tested chemical modifiers, and interestingly, the organic solvents, such as n-hexane and toluene, also favored the enzyme activity. Thus, this study involves characterization of B. amyloliquefaciens lipase at molecular level. The key outcomes are novelty of the bacterial source and purification of the enzyme with desirable properties for industrial applications.
Subject(s)
Adult , Female , Humans , Male , Diet/psychology , Environment Design , Food Supply/methods , Obesity/epidemiology , Residence Characteristics/statistics & numerical data , Body Mass Index , Beverages/statistics & numerical data , Commerce , Diet/ethnology , Diet/statistics & numerical data , Energy Intake , Fast Foods , Fruit , Food Supply/statistics & numerical data , Health Surveys , Los Angeles/epidemiology , Motor Activity , Obesity/prevention & control , Socioeconomic Factors , Surveys and Questionnaires , Sedentary Behavior/ethnology , Sweetening Agents/administration & dosage , Vegetables , Walking/statistics & numerical dataABSTRACT
Abstract Lipases are enzymes of immense industrial relevance, and, therefore, are being intensely investigated. In an attempt to characterize lipases at molecular level from novel sources, a lipase gene from Bacillus amyloliquefaciens PS35 was cloned, heterologously expressed in Escherichia coli DH5α cells and sequenced. It showed up to 98% homology with other lipase sequences in the NCBI database. The recombinant enzyme was then purified from E. coli culture, resulting in a 19.41-fold purification with 9.7% yield. It displayed a preference for long-chain para-nitrophenyl esters, a characteristic that is typical of true lipases. Its optimum pH and temperature were determined to be 8.0 and 40 °C, respectively. The half-lives were 2.0, 1.0 and 0.5 h at 50 °C, 60 °C and 70 °C, respectively. The metal ions K+ and Fe3+ enhanced the enzyme activity. The enzyme displayed substantial residual activity in the presence of various tested chemical modifiers, and interestingly, the organic solvents, such as n-hexane and toluene, also favored the enzyme activity. Thus, this study involves characterization of B. amyloliquefaciens lipase at molecular level. The key outcomes are novelty of the bacterial source and purification of the enzyme with desirable properties for industrial applications.(AU)