Autodisplay for the co-expression of lipase and foldase on the surface of E. coli: washing with designer bugs.
BACKGROUND
Lipases including the lipase from Burkholderia cepacia are in a main focus in biotechnology research since many years because of their manifold possibilities for application in industrial processes. The application of Burkholderia cepacia lipase for these processes appears complicated because of the need for support by a chaperone, the lipase specific foldase. Purification and reconstitution protocols therefore interfere with an economic implementation of such enzymes in industry. Autodisplay is a convenient method to express a variety of passenger proteins on the surface of E. coli. This method makes subsequent purification steps to obtain the protein of interest unnecessary. If enzymes are used as passengers, the corresponding cells can simply be applied as whole cell biocatalysts. Furthermore, enzymes surface displayed in this manner often acquire stabilization by anchoring within the outer membrane of E. coli.


RESULTS
The lipase and its chaperone foldase from B. cepacia were co-expressed on the surface of E. coli via autodisplay. The whole cell biocatalyst obtained thereby exhibited an enzymatic activity of 2.73 mU mL⁻¹ towards the substrate p-nitrophenyl palmitate when applied in an OD₅₇₈ =1. Outer membrane fractions prepared from the same culture volume showed a lipase activity of 4.01 mU mL⁻¹. The lipase-whole cell biocatalyst as well as outer membrane preparations thereof were used in a standardized laundry test, usually adopted to determine the power of washing agents. In this test, the lipase whole cell biocatalyst and the membrane preparation derived thereof exhibited the same lipolytic activity as the purified lipase from B. cepacia and a lipase preparation which is already applied in commercial washing agents.


CONCLUSIONS
Co-expression of both the lipase and its chaperone foldase on the surface of E. coli yields a lipid degrading whole cell biocatalyst. Therefore the chaperone supported folding process, absolutely required for the lipolytic activity appears not to be hindered by surface display. Furthermore, the cells and the membrane preparations appeared to be stable enough to endure a European standard laundry test and show efficient fat removal properties herein.
Eva
E
Kranen
Christian
C
Detzel
Thomas
T
Weber
Institute of Pharmaceutical and Medicinal Chemistry, PharmaCampus, Westfalian Wilhelms-University Münster, Corrensstr, 48, 48149 Münster, Germany. joachim.jose@uni-muenster.de.
Joachim
J
Jose
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D016956:Burkholderia cepacia / Q000201:enzymology / Q000235:genetics; D002462:Cell Membrane / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D008049:Lipase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010168:Palmitates / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
D001426:Bacterial Proteins; D018832:Molecular Chaperones; D010168:Palmitates; D011993:Recombinant Fusion Proteins; C050015:4-nitrophenyl palmitate; D008049:Lipase