Epsilon Open Archive

Abstract

Adhesion is an important first step in infection, where the microorganism attaches to a host cell. In many cases adhesion is mediated by fimbriae, or pili; hairlike organelles composed of a large number of protein subunits, protruding out from the bacterial surface. In this thesis, the adhesion and assembly of two such organelles has been studied: type-1 pili from Escherichia coli and the capsular F1 antigen from Yersinia pestis. The adhesin molecule of type-1 pili is FimH, and the structure of the FimH lectin domain was determined to 2.3 Å. High structural similarity to the same domain in the FimH:FimC adhesin:chaperone structure shows rigidity and structural independency of the lectin domain. In the crystal structure a butyl mannoside was discovered in the FimH binding site. Binding studies of alkyl mannosides and aryl mannoside show that E. coli FimH recognizes these two classes of compounds with high affinity. Using a series of trimannosides corresponding to structures present in N-linked high-mannose glycoproteins, the binding properties of FimH from two different UPEC and one fecal E. coli strains were investigated. Our results suggest that the differences in adhesion phenotype mediated by these different adhesins are caused by differences in adhesin presentation rather than by affinity differences. The antiphagocytic capsule around Yersinia pestis is constructed from multiple copies of the Caf1 subunit assembled into thin fibres. The structure of the Caf1M:Caf1 chaperone:subunit binary complex and the Caf1M:Caf1:Caf1 ternary complex from Y. pestis was determined. Comparison of the chaperone bound Caf1 subunit with the Caf1 fibre subunit revealed that the Caf1M chaperone jams the folding of Caf1 in a high-energy conformation with a poorly packed hydrophobic core. When the chaperone dissociates and is replaced by the donor strand from the next subunit, folding is allowed to continue to completion. The folding energy released in this step is proposed to drive fibre assembly.