Engineering of a Staphylococcus carnosus surface display system by substitution or deletion of a Staphylococcus hyicus lipase propeptide.
Surface display of recombinant proteins on bacteria and phages has become an important topic in bioscience. A system for the display of heterologous proteins on the surface of Staphylococcus carnosus employs the secretion signal and propeptide from a Staphylococcus hyicus lipase for translocation and since the propeptide is of considerable size (207 amino acids) and not processed in S. carnosus, we have investigated the possibility to delete or substitute the propeptide for smaller protein domains, to thereby improve the surface display system. A set of new vectors was constructed and the surface expression of model proteins was investigated by various methods, including fluorescence-activated cell sorting. The results suggest that the propeptide region indeed can be deleted when proteins which are easily secretable are displayed. In contrast, the propeptide seems to be advantageous for translocation of inefficiently secreted proteins. Moreover, our study also presents a rational strategy for how to monitor the engineering efforts for the optimization of a surface display system.
Samuelson
P
P
Department of Biotechnology, Kungliga Tekniska Högskolan, S-100 44, Stockholm, Sweden.
Cano
F
F
Robert
A
A
Ståhl
S
S
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D017931:DNA Primers; D004792:Enzyme Precursors / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli; D005818:Genetic Engineering; D008049:Lipase / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D021382:Protein Sorting Signals / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D013210:Staphylococcus / Q000201:enzymology* / Q000235:genetics*
D017931:DNA Primers; D004792:Enzyme Precursors; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D008049:Lipase