초록
<P><B>Background</B></P><P>Bacteria included in the genus <I>Streptomyces</I> exhibit several attractive characteristics that make them adequate hosts for the heterologous expression of proteins. One of them is that some of its species have a high secretion capacity and hence the protein of interest could be released to the culture supernatant, facilitating downstream processing. To date, all the expression vectors described for these bacteria contain antibiotic resistance genes as selection markers. However, the use of antibiotics to produce proteins at industrial level is currently becoming more restricted owing to the possibility of contamination of the final product. In this report, we describe the use of the <I>S. lividans yefM/yoeBsl</I> toxin-antitoxin system to develop a stable plasmid expression system.</P><P><B>Results</B></P><P>In order to use the <I>yefM/yoeBsl</I> system to stabilize expression plasmids in <I>Streptomyces</I>, a <I>S. lividans</I> mutant strain that contained only the toxin gene (<I>yoeBsl</I>) in its genome and the antitoxin gene (<I>yefMsl</I>) located in a temperature-sensitive plasmid was constructed and used as host. This strain was transformed with an expression plasmid harbouring both the antitoxin gene and the gene encoding the protein of interest. Thus, after elimination of the temperature-sensitive plasmid, only cells with the expression plasmid were able to survive. On using this system, two proteins - an α-amylase from <I>S. griseus</I> and a xylanase from <I>S. halstedii -</I> were overproduced without the addition of antibiotic to the culture medium. The production of both proteins was high, even after long incubations (8 days), and after serial subcultures, confirming the stability of the plasmids without antibiotic selection.</P><P><B>Conclusions</B></P><P>This is the first report that describes the use of a toxin-antitoxin system to maintain high -copy plasmids in <I>Streptomyces</I>. This finding could be a valuable tool for using <I>Streptomyces</I> as a host to produce proteins at the industrial and pharmaceutical levels without the use of antibiotics in the production step.</P>