Chemical Biology Reveals Involvement of N-acetylglucosaminidase LytG In Cell Elongation and Division.

The growth and division of the Gram-positive cell requires the coordinated action of enzymes involved in the synthesis and degradation of the heteropolymer peptidoglycan. Herein, we present the use of the diamide Masarimycin, an inhibitor of the exo-N-acetylglucosaminidase (GlcNAcase) LytG, as a chemical biology probe to elucidate biological role of this cell wall degrading enzyme. Using a combination of chemical biology and genetic approaches we provide the first evidence that LytG activity influences the elongation and division complexes in Bacillus subtilis. Chemical inhibition of LytG results in dysregulated cell elongation and localization of division plane and the induction of the cell wall stress response. In the presence of masarimycin, cells are elongated 3-fold compared to wild-type cells and formation of daughter cells along the sidewall was observed. The use of genetic and synergy/antagonism screens established connections to late-stage peptidoglycan synthesis, particularly related to cross-linking function. These results stand in stark contrast to those observed for the lytG knockout, which does not exhibit these phenotypes. The results presented here emphasize the difficulty in assigning mode-of-action using solely genetic screens. This is the first report of assigning physiological role of LytG beyond being the major GlcNAcase during vegetative growth.