초록
<P>Riboswitches are natural RNA elements that regulate gene expression by binding a ligand. Here, we demonstrate the possibility of altering a natural lysine-OFF riboswitch from <I>Eschericia coli</I> (ECRS) to a synthetic lysine-ON riboswitch and using it for metabolic control. To this end, a lysine-ON riboswitch library was constructed using <I>tetA</I>-based dual genetic selection. After screening the library, the functionality of the selected lysine-ON riboswitches was examined using a report gene, <I>lacZ</I>. Selected lysine-ON riboswitches were introduced into the <I>lysE</I> gene (encoding a lysine transport protein) of <I>Corynebacterium glutamicum</I> and used to achieve dynamic control of lysine transport in a recombinant lysine-producing strain, <I>C. glutamicum</I> LPECRS, which bears a deregulated aspartokinase and a lysine-OFF riboswitch for dynamic control of the enzyme citrate synthase. Batch fermentation results of the strains showed that the <I>C. glutamicum</I> LPECRS strain with an additional lysine-ON riboswitch for the control of <I>lysE</I> achieved a 21% increase in the yield of lysine compared to that of the <I>C. glutamicum</I> LPECRS strain and even a 89% increase in yield compared to that of the strain with deregulated aspartokinase. This work provides a useful approach to generate lysine-ON riboswitches for <I>C. glutamicum</I> metabolic engineering and demonstrates for the first time a synergetic effect of lysine-ON and -OFF riboswitches for improving lysine production in this industrially important microorganism. The approach can be used to dynamically control other genes and can be applied to other microorganisms.</P><P><B>Graphic Abstract</B><BR><IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/asbcd6/2015/asbcd6.2015.4.issue-12/acssynbio.5b00075/production/images/medium/sb-2015-00075v_0004.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/sb5b00075'>ACS Electronic Supporting Info</A></P>