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One-step purification and immobilization of his-tagged GL-7-ACA acylase

Paper ID Volume ID Publish Year Pages File Format Full-Text
17897 42704 2007 6 PDF Available
Title
One-step purification and immobilization of his-tagged GL-7-ACA acylase
Abstract

GL-7-ACA acylase (GLA) is an enzyme that converts GL-7-ACA to 7-ACA, a starting material for semi-synthetic cephalosporin antibiotics. Four his-tagged GLAs were constructed in this work different in their N-terminal amino acid residues. Among them the highest activity of GLA reached 3800 U/l in 19 h for the recombinant strain BL21 (DE3)/pET28GA01. The immobilized GLA on TALON affinity support, agar-EPI-IDA-Co2+ and FP-IDA-Ni2+ was evaluated by one-step purification and immobilization, and the highest immobilized activity was 163 U/g for TALON affinity support. But more robust matrix made FP-IDA-Ni2+ suitable for the industrial application. The activities of four immobilized GLAs on FP-IDA-Ni2+ ranged from 66.7 to 80 U/g. Immobilized GLA showed good stability at pH value below 11.0 and at temperature up to 30 °C. To the consecutive conversion of GL-7-ACA, immobilized GLA was recycled 21 times without significant loss of activity, and the average yield rate of 7-ACA reached 90%. These results indicated that intracellular production of his-tagged GLA in E. coli, followed by one-step affinity purification and immobilization on FP-IDA-Ni2+ resins, might serve as an effective process for the industrial application.

Keywords
CPC, cephalosporin C; GL-7-ACA, glutaryl-7-aminocephalosporanic acid; 7-ACA, 7-aminocephalosporanic acid; GLA, GL-7-ACA acylase; CA, cephalosporin acylase; BV/min, bed volume per minute; Ntn, N-terminal nucleophile; vvm, volume of gas per volume of liquid
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One-step purification and immobilization of his-tagged GL-7-ACA acylase
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 41, Issue 4, 3 September 2007, Pages 474–479
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