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Efficient cloning and expression of a thermostable nitrile hydratase in Escherichia coli using an auto-induction fed-batch strategy

Paper ID Volume ID Publish Year Pages File Format Full-Text
34422 45025 2013 7 PDF Available
Title
Efficient cloning and expression of a thermostable nitrile hydratase in Escherichia coli using an auto-induction fed-batch strategy
Abstract

•A NHase gene was functionally expressed in E. coli assisted by an activator gene.•The NHase exhibited adequate thermal stability.•The NHase can efficiently hydrate 3-cyanopyridine to produce nicotinamide.•The recombinant NHase was efficiently produced using an auto-induction strategy.

A nitrile hydratase (NHase) gene from Aurantimonas manganoxydans was cloned and expressed in Escherichia coli BL21 (DE3). A downstream gene adjacent to the β-subunit was necessary for the functional expression of the recombinant NHase. The structural gene order of the Co-type NHase was α-subunit beyond β-subunit, different from the order typically reported for Co-type NHase genes. The NHase exhibited adequate thermal stability, with a half-life of 1.5 h at 50 °C. The NHase efficiently hydrated 3-cyanopyridine to produce nicotinamide. In a 1-L reaction mixture, 3.6 mol of 3-cyanopyridine was completely converted to nicotinamide in four feedings, exhibiting a productivity of 187 g nicotinamide/g dry cell weight/h. An industrial auto-induction medium was applied to produce the recombinant NHase in 10-L fermenter. A glycerol-limited feeding method was performed, and a final activity of 2170 U/mL culture was achieved. These results suggested that the recombinant NHase was efficiently cloned and produced in E. coli.

Keywords
Nitrile hydratase; Thermostable; Recombinant expression; Escherichia coli; Auto-induction; Fed-batch cultivation
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Efficient cloning and expression of a thermostable nitrile hydratase in Escherichia coli using an auto-induction fed-batch strategy
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Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 48, Issue 12, December 2013, Pages 1921–1927
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
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Don't Miss Today's Special Offer
Price was $35.95
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Price after discount Only $4.95
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