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Extracellular expression of Thermobifida fusca cutinase with pelB signal peptide depends on more than type II secretion pathway in Escherichia coli

Paper ID Volume ID Publish Year Pages File Format Full-Text
22804 43392 2015 6 PDF Available
Title
Extracellular expression of Thermobifida fusca cutinase with pelB signal peptide depends on more than type II secretion pathway in Escherichia coli
Abstract

•The pelB-cutinase was extracellularly expressed in Escherichia coli.•The periplasmic and cytoplasmic fractions had phospholipid hydrolase activity.•The recombinant cells showed improved membrane permeability.•The secretion efficiency of inactive mutant significantly decreased.•The activity of pelB-cutinase plays a main role in its extracellular production.

Our previous studies demonstrated that Thermobifida fusca cutinase is released into culture medium when expressed without a signal peptide in Escherichia coli, and this extracellular expression results from an enhanced membrane permeability caused by cutinase's phospholipid hydrolase activity. The present study investigated whether this phenomenon would also occur during the expression of cutinase fused to pelB signal peptide (pelB-cutinase). Secretion of fusion proteins of this type is generally believed to occur via type II secretion pathway. The results showed that when pelB-cutinase was expressed in a secB knockout strain, which has a defective type II secretion pathway, there was still a large amount of cutinase in the culture medium. Additional experiments confirmed that the periplasmic and cytoplasmic fractions of the expressing cells had hydrolytic activity toward phosphatidyl ethanolamine, and the recombinant cells showed correspondingly improved membrane permeability. All these phenomena were also observed in the parent E. coli strain. Moreover, the secretion efficiency of the inactive cutinase mutant was found to be significantly lower than that of pelB-cutinase in the parent E. coli. Based on these results, the phospholipid hydrolase activity of pelB-cutinase must play a larger role in its extracellular production than does type II secretion pathway.

Keywords
Thermobifida fusca cutinase; Escherichia coli; pelB signal peptide; Extracellular expression; Phospholipid hydrolase activity
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Extracellular expression of Thermobifida fusca cutinase with pelB signal peptide depends on more than type II secretion pathway in Escherichia coli
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 204, 20 June 2015, Pages 47–52
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