Effective application of whole-cell devices in synthetic biology and biocatalysis will always require
consideration of the uptake of molecules of interest into the cell. Here we demonstrate that the AlkL protein
from Pseudomonas putida GPo1 is an alkane import protein capable of industrially relevant rates of uptake
of C7-C16 n-alkanes. Without alkL expression, native E.coli n-alkane uptake was the rate-limiting step in
both the whole-cell bioconversion of C7-C16 n-alkanes and in the activation of a whole-cell alkane biosensor
by C10 and C11 alkanes. By coexpression of alkL as a transporter plug-in, specific yields improved by up to
100-fold for bioxidation of>C12 alkanes to fatty alcohols and acids. The alkL protein was shown to be toxic
to the host when overexpressed but when expressed from a vector capable of controlled induction, yields of
alkane oxidation were improved a further 10-fold (8 g/L and 1.7 g/g of total oxidized products). Further
testing of activity on n-octane with the controlled expression vector revealed the highest reported rates of
120 μmol/min/g and 1 g/L/h total oxidized products. This is the first time AlkL has been shown to directly
facilitate enhanced uptake of C10-C16 alkanes and represents the highest reported gain in product yields
resulting from its use.
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