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Production of d-hydantoinase via surface display and self-cleavage system

Paper ID Volume ID Publish Year Pages File Format Full-Text
20552 43180 2013 5 PDF Available
Title
Production of d-hydantoinase via surface display and self-cleavage system
Abstract

In this study, the cell surface expression system was the first time used to directly produce extracellular enzyme. In the plasmid construction, the truncated ice nucleation protein (INP) was fused with intein (INT) and target protein, d-hydantoinase (DHTase), to form the INP-INT-DHTase gene. The plasmid containing this gene was transformed into Escherichia coli ER2566 cells. The gene construct enables the expression of INP-INT-DHTase fusion protein, which might anchor on cell membrane surface. The induction conditions were studied and optimal conditions were as follows: E. coli ER2566 was incubated at 37°C and 200 rpm till OD600 reached 0.6. Then, 0.05 mM IPTG was added and the induction was conducted at 15°C for 24 h. The cell was harvested and resuspended in the cleavage buffer (50 mM Tris-HCl buffer, pH 6). The cleavage reaction was carried out at 25°C, and 100 rpm for 24 h. The DHTase with an activity of 0.225 U/ml and a purity of 63.2% was obtained via centrifugation. This study demonstrated the feasibility of direct extracellular enzyme production using E. coli via only two steps of centrifugation.

Keywords
Biocatalysis; Enzyme production; Ice nucleation protein-intein surface display system; Protein recovery; Self-cleavage; d-Hydantoinase
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Production of d-hydantoinase via surface display and self-cleavage system
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Bioscience and Bioengineering - Volume 116, Issue 5, November 2013, Pages 562–566
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us