| The focus of this dissertation is structural evaluation of proteins. The first three chapters relate to the proteins of the bacterial flagellar motor. The final four chapters are pertinent to an enzyme within the porphyrin biosynthetic pathway. I have used the tools of molecular biology, biochemistry, and x-ray crystallography to probe these proteins.; Through the design and cloning of mutants, as well as binding studies between mutant proteins, I have mapped the regions of the protein FliM that are important in binding to the other flagellar proteins FliG, FliN, and CheY. Analytical ultra-centrifugation has been used to look at the possible solution states of FliM and FliN and perhaps shed some light on their states before being incorporated into the motor. X-ray protein crystallography is being attempted to determine the atomic structure of several of these flagellar proteins to better understand their function and mechanism of action.; X-ray crystallography was used to determine the atomic structure of the fourth enzyme in the porphyrin biosynthetic pathway, uroporphyrinogen III synthase. An introduction to this enzyme is presented in Chapter 4, an overview of the method of x-ray crystallography comprises Chapter 5, the structure and discussion of the enzyme is in Chapter 6, and future directions for this research are found in Chapter 7. |
