NDLI: Outer Membrane Protein AlkL Boosts Biocatalytic Oxyfunctionalization of Hydrophobic Substrates in Escherichia coli

Content Provider	PubMed Central
Author	Julsing, Mattijs K. Manfred, Schrewe Cornelissen, Sjef Hermann, Inna Schmid, Andreas Bühler, Bruno
Copyright Year	2012
Abstract	The outer membrane of microbial cells forms an effective barrier for hydrophobic compounds, potentially causing an uptake limitation for hydrophobic substrates. Low bioconversion activities (1.9 U gcdw −1) have been observed for the ω-oxyfunctionalization of dodecanoic acid methyl ester by recombinant Escherichia coli containing the alkane monooxygenase AlkBGT of Pseudomonas putida GPo1. Using fatty acid methyl ester oxygenation as the model reaction, this study investigated strategies to improve bacterial uptake of hydrophobic substrates. Admixture of surfactants and cosolvents to improve substrate solubilization did not result in increased oxygenation rates. Addition of EDTA increased the initial dodecanoic acid methyl ester oxygenation activity 2.8-fold. The use of recombinant Pseudomonas fluorescens CHA0 instead of E. coli resulted in a similar activity increase. However, substrate mass transfer into cells was still found to be limiting. Remarkably, the coexpression of the alkL gene of P. putida GPo1 encoding an outer membrane protein with so-far-unknown function increased the dodecanoic acid methyl ester oxygenation activity of recombinant E. coli 28-fold. In a two-liquid-phase bioreactor setup, a 62-fold increase to a maximal activity of 87 U gcdw −1 was achieved, enabling the accumulation of high titers of terminally oxyfunctionalized products. Coexpression of alkL also increased oxygenation activities toward the natural AlkBGT substrates octane and nonane, showing for the first time clear evidence for a prominent role of AlkL in alkane degradation. This study demonstrates that AlkL is an efficient tool to boost productivities of whole-cell biotransformations involving hydrophobic aliphatic substrates and thus has potential for broad applicability.
Related Links	http://dx.doi.org/10.1128/aem.00949-12
Ending Page	5733
Page Count	10
Starting Page	5724
File Format	PDF
ISSN	00992240
e-ISSN	10985336
Journal	Applied and Environmental Microbiology
Issue Number	16
Volume Number	78
Language	English
Publisher	American Society for Microbiology
Publisher Date	2012-08-15
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Biotechnology Food Science Ecology Applied Microbiology and Biotechnology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Ecology Food Science Applied Microbiology and Biotechnology Biotechnology

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