Characterization of the adhesive holdfast of marine and freshwater caulobacters

Caulobacters are prosthecate (stalked) bacteria that elaborate an attachment organelle called a holdfast at the tip of the cellular stalk. We examined the binding of lectins to the holdfasts of 16 marine Caulobacter strains and 10 freshwater species or strains by using a panel of fluorescein-conjugated lectins and fluorescence microscopy. The holdfasts of all the marine isolates bound to only wheat germ agglutinin (WGA) and other lectins that bind N-acetylglucosamine (GlcNac) residues. The freshwater caulobacters showed more variability in holdfast composition. Some bound only to WGA and comparable lectins as the marine strains did. Others bound additional or other lectins, and some did not bind to the lectins tested. The binding of WGA appeared to involve the regions of the holdfast involved with adhesion; a holdfast bound to WGA was significantly less adhesive to glass. Competition experiments with WGA-binding holdfasts and oligomers of GlcNac demonstrated that trimers of GlcNac (the preferred substrate for WGA binding) were more effective than dimers or monomers in preventing WGA binding to holdfasts, suggesting that stretches of contiguous GlcNac residues occur in the WGA-binding holdfasts. In addition, differences between freshwater and marine holdfasts in the strength of WGA binding were noted. The effect of a number of proteolytic and glycolytic enzymes on holdfast integrity was examined; the proteases had no effect for all caulobacters. None of the glycolytic enzymes had an effect on marine caulobacter holdfasts, but chitinase and lysozyme (both attack oligomers of GlcNac) disrupted the holdfasts of those freshwater caulobacters that bound WGA. Despite some similarity to chitin, holdfasts did not bind Calcofluor and no measurable effects on holdfast production were detectable after cell growth in the presence of diflubenzuron or polyoxin D, inhibitors of chitin synthesis in other systems. Finally, the holdfasts of all caulobacters bound to colloidal gold particles, without regard to the coating used to stabilize the gold particles. This binding was stronger or more specific than WGA binding; treatment with colloidal gold particles prevented WGA binding, but the reverse was not the case.