Date of Award


Document Type


Degree Name

Doctor of Philosophy (PhD)


Biological Sciences

First Advisor

Randall C. Gayda


FtsA is a gene essential for cell septation. It has been postulated to have both regulatory and structural functions. To study the structural involvement of the FtsA protein in septum formation, four FtsA-LacZ protein fusions were constructed using a mini-Mu transposon (MudII1734). When maxicells containing either ftsA-lacZ or ftsA specifying plasmids were radioactively labelled and fractionated, FtsA-LacZ fusion protein and FtsA were located in both the membrane and cytoplasmic fractions. The FtsA-LacZ fusion proteins were also located in dividing wild type dividing cells by measuring $\beta$-galactosidase activity. Cell envelope fractions from a sucrose equilibrium gradient had $\beta$-galactosidase activity in outer membrane-heavy(OM$\sb{\rm H}$) and outer membrane-light (OM$\sb{\rm L}$) fractions. To verify the cellular location of native FtsA protein in membrane fractions, an expression plasmid was constructed by placing a ftsA-lacZ gene fusion downstream from a $\lambda$ P$\sb{\rm L}$ promoter. The FtsA-LacZ fusion protein was overproduced and purified by $\beta$-galactosidase affinity column chromatography. Polyclonal antibodies to this protein were isolated and demonstrated to be specific for the FtsA-LacZ fusion, FtsA protein and $\beta$-galactosidase. Cross reactivity to FtsA protein was found in outer membrane fractions, OM$\sb{\rm H}$ and OM$\sb{\rm L}$, and inner membrane fraction of wild type cells. These data suggest that the FtsA protein may be functioning as an inner-outer membrane protein in septation. Electron microscopy of thin sections treated with FtsA-LacZ antibodies and immunogold labelling also suggest that the FtsA protein is located in the cell envelope. Furthermore, direct identification of FtsA protein by immuno-electron-microscopy in normal and filamentous cells indicates an enrichment of FtsA protein near septation sites.