Substrate evaluation of the nitrile degrading enzymes from Rhodococcus rhodochrous ATCC BAA 870

• Main Author: Mashweu, Adelaide
• Format: Others
• Language: en
• Published: 2021
• Online Access: https://hdl.handle.net/10539/31394

A dissertation submitted to the Faculty of Science, University of the Witwatersrand in fulfilment for the requirements of the degree of Master of Science, 2020 === The focus of this research was the nitrile degrading enzymes nitrile hydratases (NHase) and whole cell nitrilases. NHases and whole cell nitrilases have not been as extensively applied in industry as they could be due to a number of reasons, including missing information relating to their substrate scope. Therefore, this research focussed on exploring the activity of both NHase and whole cell nitrilase towards a number of nitrile compounds. The Groebke-Blackburn-Bienayméreaction (GBB) and Suzuki-Miyaura coupling reaction were employed to synthesize aromatic nitrile-bearing imidazo[1,2-a]pyridine and biaryl compounds respectively. These compounds were of varying sizes and the nitrile group was subjected to different electronic effects through incorporation of different substituents. Vinyl nitrile compounds were synthesized using the Morita-Baylis Hillman reaction (MBH), while simple nitrile compounds were purchased. The synthesized nitrile-bearing imidazo[1,2-a]pyridine compounds were subjected to both NHase and whole cell nitrilase, however, these enzymes were inactive towards these compounds. NHase showed activity towards the biaryl compounds, however, there was complete loss of activity when the biaryl compounds had a 3,4-dimethoxy group as a substituent irrespective of its position relative to the nitrile group. There was also no activity when the 3,4-difluorophenyl group was in the ortho-position relative to the nitrile functional group. NHase also showed activity towards the MBH compounds, however, the rate of hydrolysis was slow in comparison with that of the biaryl compounds. NHase was inactive towards one MBH compound bearing a trimethoxyphenylgroup as a substituent. Whole cell Nitrilase had no activity towards the MBH compounds and extending the time for hydrolysis resulted in no significant changes. NHase demonstrated excellent activity towards the simple commercially available nitrile compounds, however, the rate of hydrolysis towards nitrile compounds bearing electron-withdrawing substituents was faster in comparison with those bearing electron-donating substituents === CK2021