Under construction. Last update March 28 1995

Adhesive Pili from Gram negative bacteria

Some basic definitions

Adhesin

Carbohydrate-binding moeity of adhesive pili.

Adhesive pili

Thin fibers or fibrillae used by Gram negaitve bacteria to stick to host cells and avoid clearance. Determinants of host cell specificity and tropism.

Chaperone

A protein molecule that assists other protein molecules in attaining their native structure. Here, a molecule that helps fold pilus subunits and prevents them from aggregating in the periplasm.

Pilin

Pilus subunit.

Usher

Anchors pili to the outer membrane and functions as an assembly platform for building pili from the component subunits.

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Escherichia coli can express a number of different types of pili that are usually 5-10 nm in diameter and up to 2 microns in length. Lectin-like adhesive proteins associated with these pili mediate binding to specific carbohydrate moieties of glycolipids or glycoproteins such as alpha-mannose (type 1 pili), alpha- galactosyl-1,4-beta-galactose (P pili), alpha-sialyl-2,3-beta-galactose (S pili), or galactose/ N-acetylgalactoseamine/serin (M pili).

Type 1 and P pili have been analyzed in some detail. These pili are complex structures composed of several different protein subunits that require specific pilus-assembly machinery for the incorporation of the component subunits into functional pili. A periplasmic chaperone, and an outer membrane protein called an usher, act in concert to regulate the correct protein-protein interactions required for pilus assembly. Many other analogous pili systems have been identified in a number of different pathogenic bacterial strains, including Salmonella typhimurium, Salmonella enteriditis, Bortadella pertussis, Yersina pestis, Klebsiella Pneumoniae, and Haemophilus influenzae.

The major component of type 1 pili is repeating FimA subunits arranged in a right-handed helix to form a 7-nm-wide fiber with an axial hole. FimF, FimG, and the mannose-binding adhesin FimH are three minor components of type 1 pili. FimA, FimF, and FimG, are all about 140 amino acids long and have homologous amino acid sequences, in particular at the C-terminus. The FimH adhesin is approximately twice as large and is divided in two domains, one receptor-binding domain, and one chaperone- binding domain that is homologous to FimA, FimF, and FimG. A membrane bound protein, FimD, serves to anchor the pilus to the outer membrane of the bacteria, and also functions as the initiation site and assembly platform for pilus formation. A periplasmic chaperone protein, FimC, is required for correct folding of the subunits, and for preventing the formation of non-productive, insoluble, complexes between the different components of the pilus by forming stable, discrete, and soluble pre-assembly complexes with the pili proteins in the periplasm. Chaperone/ subunit pre-assembly complexes are targeted to the outer membrane usher where the subunits are incorporated into the growing pilus in a defined order.

The structure of the P-pilus chaperone, PapD, has previously been solved by Holmgren & Branden (1989). Pilus chaperones from other pili systems show about 30% identity to PapD and are thus all expected to have the same general fold as PapD. The 218 amino acids in the PapD polypeptide chain are folded as two immunoglobulin domains giving the molecule the overall shape of a boomerang, with a deep cleft between the domains. On the basis of the clustering of a number of highly conserved residues in this cleft, as well as on peptide-binding studies, this cleft has been proposed to be the binding site for pili proteins (Kuehn et al., 1993).

We are currently studying chaperones from five different pili systems.

A number of pre-assembly complexes have been purified by Scott Hultgrens group in St. Louis. Two such complexes are currently being used for crystallography.

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