Abstract
M.Sc. (Biochemistry)
The use of enzymes as biocatalysts in industrial processes has prompted the search for novel biocatalysts with improved properties. Carboxyl esterases are amongst the most used biocatalysts in various industrial processes, including pharmaceutical, food and textile industries. The most attractive properties of these enzymes are the high regio- and stereospecificity and stability and activity in organic solvent. However, natural enzymes are poorly suited for industrial applications that often occur under extreme conditions such as elevated temperature. As a result, there is a need to optimize/improve properties of enzymes prior to application at industrial scale. Protein engineering is a technique that was introduced in order to study structure-function properties of enzymes and (or) to design mutant enzymes with new or desirable properties.
In a previous study, three carboxyl esterase-encoding genes from selected Bacillus species were previously cloned, functionally expressed in Escherichia coli DE3 (BL21) and their kinetic data recorded. Carboxylesterases from Bacillus pumilus and Bacillus licheniformis showed the highest activity at 45 °C and pH 7.5 and 8.0, respectively. Both enzymes lost more than 60% residual activity after 10 min incubation at temperatures above 50 °C. On the other hand, a carboxyl esterase from Geobacillus kaustophilus was observed to be relatively stable with the highest activity recorded at 60 °C and pH 8.0.
When the amino acid sequences of the above-mentioned enzymes and selected members of the family VII of lipolytic enzymes were aligned it was observed that, the N-terminal regions were relatively conserved while C-terminal regions were variable. These observations formed the basis of the current study which is aimed...