Determining the crystal structure of SseB, a product of the Salmonella Pathogenicity Island II Type III Secretion System of Salmonella typhimurium
Salmonella typhimurium is a bacterium that causes many food-borne illnesses such as gastrointestinal infections, diarrhea, and abdominal cramps. It affects 700,000 to 3.8 million people each year. The SseB protein, a part of the Salmonella Pathogenicity Island II (SPI2), plays a critical role in the pathogenesis of gastrointestinal infections. It is part of the Type III Secretion System (TTSS) that is involved in translocating proteins from the bacteria to the host gastro intestinal-epithelial cells. The aim of the research project is to clone, purify the SseB protein from Salmonella typhimurium and obtain a diffracting-quality crystal that will give high resolution data so that the structure of the protein can be determined using x-ray diffraction patterns.
The SseB gene was amplified and cloned into pET30b vector and transformed into E. coli novablue cells. The SseB protein is then expressed into E. coli BL21 cells and purified using various chromatographic methods. Purified protein was used for crystallization and diffracting quality crystals were obtained using grid screening method. SseB protein crystallized in P-orthorhombic space group (P 21 21 21) and diffracted to 3.8Å. Further optimization is underway to get a good diffracting quality crystal.
It is important to determine the crystal structure of SseB since this will reveal its role in the interaction with other translocation complex such as (SseC and SseD). Based on the structural information, potential drug targets can be designed for translocon complex, which can further prevent diseases caused by the bacterium Salmonella and other closely related bacteria.
School:Texas A&M University
School Location:USA - Texas
Source Type:Master's Thesis
Keywords:translocating proteins gastrointestinal infections food borne illnesses drug target crystal structure salmonella typhimurium type iii secretion system pathogenicity island ii sseb
Date of Publication:03/20/2008