Development of biocatalyst by engineering nitrile reducing enzymes.

• Main Author: Chua, Ming Boon.
• Other Authors: Liang Zhao-Xun
• Format: Final Year Project
• Language: English
• Published: 2012
• Subjects: DRNTU::Science::Biological sciences::Biochemistry
• Online Access: http://hdl.handle.net/10356/49252
• Institution: Nanyang Technological University

Enzymes have been extensively utilized as biocatalysts in many industrial processes, ranging from the bulk manufacturing of value-added products to providing of services such as bioremediation. While the use of enzymes spans many applications, its biocatalytic role in the pharmaceutical industry has been the most significant. QueF, an oxidoreductase which catalyses the reduction of a nitrile functional group to a primary amine, was identified recently. With nitrile reduction being an expanding chemistry in the pharmaceutical industry, QueF has immense potential to be utilized as a biocatalyst to replace conventional chemical catalysts for the reduction of nitrile-containing drug intermediates. As such, a semi-rational protein engineering approach was adopted to create a QueF mutant with nitrile oxidoreductase activity for nitrile-containing unnatural substrates. A total of eight QueF mutants were created in this project. Preliminary results from NADPH depletion assay showed that none of the mutants possess activity for the unnatural substrates. Even though differential scanning fluorimetry (DSF) detected no binding interactions between the mutants and substrates, it revealed the remarkable protein stability of the QueF mutants which is promising for future protein engineering.