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Heterologous production of Escherichia coli penicillin G acylase in Pseudomonas aeruginosa

Paper ID Volume ID Publish Year Pages File Format Full-Text
24547 43525 2009 9 PDF Available
Title
Heterologous production of Escherichia coli penicillin G acylase in Pseudomonas aeruginosa
Abstract

Penicillin G acylase (PGA) is a widely studied bacterial enzyme of great industrial importance. Since its overproduction in the original organisms is mostly limited to the intracellular bacterial spaces which may lead to aggregation and cell toxicity, we have set out to explore the host organism Pseudomonas aeruginosa that possesses the Xcp machinery for secretion of folded proteins to the extracellular medium. We have made fusion proteins, consisting of Pseudomonas Sec- or Tat-specific signal peptides, the elastase propeptide and the mature penicillin G acylase. With all constructs we obtained production of PGA in P. aeruginosa, but we observed that processing of the PGA was temperature dependent and that the active enzyme could only be found after growth at 25 °C or lower temperatures. Remarkably, the mature protein, expressed from a TatProPGA hybrid, was not only found in the extracellular medium and the periplasm, but also in the cytoplasm as assessed by comparison to the reporter beta-lactamase protein. The unusual cytoplasmic localization of the mature protein strongly suggests that processing of PGA can also occur in the cytoplasm of P. aeruginosa. The extracellular localization of the TatProPGA hybrid was found not to be dependent on the tatABC-genes. The elastase signal sequence/propeptide combination appeared to be an inadequate carrier for transporting penicillin G acylase across the outer membrane of P. aeruginosa.

Keywords
Penicillin G acylase; Pseudomonas; Type II pathway; Secretion; Propeptide
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Heterologous production of Escherichia coli penicillin G acylase in Pseudomonas aeruginosa
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 142, Issues 3–4, 15 July 2009, Pages 250–258
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