Expression and Purification of Soluble Fraction of ?-Amylase BaqA from Bacillus aquimaris MKSC 6.2
?-Amylase is one of the most widely used enzymes for industries. This enzyme is able to hydrolize starch at ?-1,4 glycosidic bonds randomly, generating oligosaccharides, such as maltose, glucose, and ?-dextrin. ?-Amylase BaqA from Bacillus aquimaris MKSC 6.2 is a raw starch-degrading enzyme. Express...
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| Format: | Final Project |
| Language: | Indonesia |
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| Online Access: | https://digilib.itb.ac.id/gdl/view/32284 |
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| Institution: | Institut Teknologi Bandung |
| Language: | Indonesia |
| Summary: | ?-Amylase is one of the most widely used enzymes for industries. This enzyme is able to hydrolize starch at ?-1,4 glycosidic bonds randomly, generating oligosaccharides, such as maltose, glucose, and ?-dextrin. ?-Amylase BaqA from Bacillus aquimaris MKSC 6.2 is a raw starch-degrading enzyme. Expression of recombinant BaqA in Escherichia coli BL21(DE3) resulted in inclusion body hence it did not have any activity towards soluble starch and/or raw starch. The purpose of this study was to obtain soluble fractions of ?-amylase BaqA with the activity towards raw starch. To achieve the research objective, the expression host for of BaqA was changed to E. coli ArcticExpress. Expression of BaqA in E. coli ArcticExpress produced a partial soluble enzyme, as indicated by the presence of a band with an apparent molecular mass of 70 kDa on Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE). Purification by an affinity chromatography using the Nickel-Nitrilotriacetic Acid (Ni-NTA) matrix generated partially pure BaqA, as shown on a SDS-PAGE electrophoregram. This enzyme exhibited specific activity of 0.153 U/mg for the lysis solution and 1.899 U/mg for the purified enzyme. This result indicated that the BaqA purity increased twelve times. |
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