A NEW ROLE FOR PAGP IN SIGNAL TRANSDUCTION

Abstract

<p>The outer membrane (OM) of Gram-negative bacteria is not a static structure; it can be remodeled in response to environmental conditions that allow bacteria to survive and function in hostile conditions. PagP, an OM enzyme, plays a crucial role in remodeling the OM by transferring a palmitate chain from a phospholipid to the proximal glucosamine unit of lipid A, which anchors lipopolysaccharides (LPS) to the external leaflet of the OM. PagP also affects cytoplasmic events of core biosynthesis in response to OM stress. A putative catalytic triad (Asp 61, His 67 and Tyr 87) at the periplasmic interface is hypothesized to play a role in transducing the signal to influence core biosynthesis. Mutations of a triad residue (Y87F), a catalytical residue (S77A) and wild-type <em>pagP</em> have all been transduced separately to the strains having PagP in inactive (<em>imp</em>⁺) and active states (<em>imp4213</em>). Correction and validation of all the strains designed for this project has been achieved. A quantitative real-time polymerase chain reaction (qPCR) approach was applied to monitor the expression of candidate genes of PagP-mediated signalling. The decreased transcription of <em>slyB </em>in a strain containing <em>pagP </em>in comparison to the strain with deleted <em>pagP </em>indicates that PagP inhibits <em>slyB</em> transcription. The periplasmic catalytic triad of PagP appears to play a role in the repression of SlyB because the Y87F mutant also showed activated <em>slyB </em>transcription. The effect of PagP over the transcription of extracytoplasmic factor, σE, and glucosyl transferase encoded by <em>waaG </em>was analyzed and ruled out because of their stable transcription in both <em>imp</em>+ and <em>imp4213 </em>strains.</p>