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Tuesday 27 September 2016

Preparation of CLEAs of a recombinant NHase ES-NHT-118

Preparation of Cross-linked Enzyme Aggregates of Nitrile Hydratase ES-NHT-118 from E. coli by Macromolecular Cross-linking Agent

Liya Zhou, Haixia Mou, Jing Gao, Li Ma, Ying He and Yanjun Jiang



Cross-linked enzyme aggregates (CLEAs) of nitrile hydratase (NHase) ES-NHT-118 from E. coli were prepared by using ammonium sulfate as precipitating agent followed by cross-linking with dextran polyaldehyde for the first time. In this process, egg white was added as an amine source to aid formation of CLEAs. The optimal conditions of the immobilization process were determined. Michaelis constants (Km) of free NHase and NHase CLEAs were also determined. The NHase CLEAs exhibited increased stability at varied pH and temperature conditions compared to its free counterpart. When exposed to high concentrations of acrylamide, NHase CLEAs also exhibited effective catalytic activity.

 

Thursday 1 September 2016

ChromSoc nitrilase flow chemistry project 5

We have looked at the conversion of 4-cyanopyridine to isonicotinic acid using our mesoscale flow chemistry apparatus containing a nitrilase enzyme immobilized in alginate. Here is an example of our results.


Rob flowed the reaction medium through the device, and took a measurement of the amount of ammonia (using our colorimetric nitrilase assay) available after each cycle. Each cycle took about 20 minutes. It has not reached completion but seems to be a fairly robust system as far as it goes.