Interactions among the outer membrane proteins of the starch utilization system of Bacteroides thetaiotaomicron

We have studied the starch utilization system (sus) of Bacteroides thetaiotaomicron as a model for polysaccharide utilization by Bacteroides. Eight genes in the starch utilization system of B. thetaiotaomicron have been identified and the gene products encoded by susC through susG are outer membrane proteins. Our hypothesis that the complex(es) of outer membrane proteins may be needed for the multiple steps involved in starch binding, starch hydrolysis, and the translocation of the hydrolyzed starch to the periplasm. A simpler system to study interactions among these outer membrane proteins was required and therefore, I constructed a mutant strain that contained a smaller number of sus genes.; To determine which Sus proteins are essential, I eliminated the encoding genes one at a time, providing susG, a gene already known to be essential, in trans. Only three proteins (SusC, SusD, and SusG) were essential for starch utilization defining the minimal starch utilization system. The growth rate of the strain carrying the minimal system was similar to that of wild type, even though it lacked two Sus proteins (SusE and SusF). Interestingly, the starch binding activity of the strain with the minimal starch utilization system was higher than wild type. In addition, the proteolytic sensitivities of the proteins to Proteinase K were different between wild type and the strain carrying the minimal system. These differences might indicate that the topology of the Sus proteins of the minimal system differs from that of the Sus proteins of wild type. Nonetheless, the minimal system works as well or better than the natural five protein systems.; In this study, the minimal starch utilization system was used to study interactions among SusC, SusD, and SusG. To examine interactions between SusE and SusF, wild type and other mutants were used. For this study, several biochemical techniques such as formaldehyde cross-linking, affinity chromatography, proteolytic sensitivity, and native gel electrophoresis were used. These results showed that SusC and SusD interact with each other, and SusE and SusF interact with each other. However, I could not detect any evidence for interactions between SusG and any other Sus protein.