Secretion and regulation of the chitinolytic machinery in Serratia marcescens
Protein secretion is an important activity used by bacterial to sense, adapt and survive in different environments. In Gram-negative bacteria proteins destined for secretion must be translocated across both the inner and outer membranes in either a one- or a two-step process. Serratia marcescens is renowned as a prolific secretor of proteins and is one of the most effective microbes at extracellular chitin degradation. To achieve this S. marcescens secretes three chitinases (ChiA, ChiB and ChiC) as well as a chitin binding protein (CBP21) that together with the periplasmic chitobiase (Chb) form the chitinolytic machinery. Research in this group had identified two genes (chiW and chiX) encoding a holin-like protein and a putative endopeptidase, that are essential for secretion of the chitinolytic machinery. Moreover chiA was found expressed by a small subpopulation of cells and in coordination with chiX. This project used fluorescence microscopy experiments to show the co-expression of chiX with chiC alongside chitinase secretion assays that provided initial insights regarding the role of protein terminus on chitinase stability and secretion. This project also investigated the role of ChiR, a LysR-type transcriptional regulator, and Hfq, a RNA-binding protein, on Serratia chitinolytic machinery. Results revealed that Hfq plays a role on ChiR synthesis which modulates chitinase and chiWX transcription. In addition, overexpression of chiR resulted in a higher population of ChiC producer cells and further investigations revealed a plethora of ChiR target genes and binding motifs.
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