Polypeptide with lipase activity and polynucleotide encoding the polypeptide
A technology of lipase and activity, applied in hydrolytic enzymes, botany equipment and methods, biochemical equipment and methods, etc.
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Embodiment 1
[0094] Example 1 - Production of lipase variants
[0095] A plasmid containing the gene encoding the polypeptide is constructed using standard methods in the art and transformed into a suitable host cell.
[0096] Fermentations were performed as fed-batch fermentations using a constant medium temperature of 34°C and an initial volume of 1.2 liters. The initial pH of the medium was set to 6.5. Once the pH increased to 7.0, by adding 10% H 3 PO 4 to keep this value. Dissolved oxygen levels in the medium were controlled by varying the rate of agitation and using a fixed aeration rate of 1.0 liters of air per liter of medium per minute. The feed addition rate was kept at a constant level throughout the fed-batch phase.
[0097] The batch medium contains maltose syrup as carbon source, urea and yeast extract as nitrogen source, and a mixture of trace metals and salts. Feeds added continuously during the fed-batch phase contained maltose syrup as a carbon source, while yeast e...
Embodiment 2
[0099] Example 2 - Lipase activity units (LU) relative to absorbance at 280 nm (LU / A280)
[0100] Lipase activity (LU) was determined as described above in the lipase activity section. The absorbance at 280 nm (A280) of the lipase was measured. The specific activity of a polypeptide can be expressed as a ratio of LU / A280.
[0101] Relative LU / A280 is calculated by dividing the LU / A280 of the polypeptide by the LU / A280 of the reference enzyme. In the context of the present invention, said reference enzyme is the lipase of SEQ ID NO: 2 with the substitution T231R+N223R.
Embodiment 3
[0102] Example 3 - Calculation of Relative Performance (RP) from Data Acquired by Automated Mechanical Stress Test (AMSA)
[0103] The polypeptides of the invention were tested using the Automatic Mechanical Stress Assay (AMSA). Using the AMSA test, it is possible to examine the wash performance of large quantities of small volume enzyme detergent solutions. The AMSA plate has multiple slots for the test solution, and a cover that compresses the textile sample to be laundered against all slot openings. During the wash time, the plate, test solution, textile and cover are shaken vigorously to bring the test solution into contact with the textile and to apply mechanical stress. For further description see WO02 / 42740, especially the paragraph "Special method embodiments" on pages 23-24. The container containing the detergent test solution consists of a cylindrical hole (diameter 6 mm, depth 10 mm) in a metal plate. The soiled fabric (test material) was placed on top of the met...
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