Robust cross-links in molluscan adhesive gels: Testing for contributions from hydrophobic and electrostatic interactions

Abstract

The cross-linking interactions that provide cohesive strength to molluscan adhesive gels were investigated. Metal-based interactions have been shown to play an important role in the glue of the slug Arion subfuscus (Draparnaud), but other types of interactions may also contribute to the glue's strength and their role has not been investigated. This study shows that treatments that normally disrupt hydrophobic or electrostatic interactions have little to no effect on the slug glue. High salt concentrations and non-ionic detergent do not affect the solubility of the proteins in the glue or the ability of the glue proteins to stiffen gels. In contrast, metal chelation markedly disrupts the gel. Experiments with gel filtration chromatography identify a 40 kDa protein that is a central component of the cross-links in the glue. This 40 kDa protein forms robust macromolecular aggregations that are stable even in the presence of high concentrations of salt, non-ionic detergent, urea or metal chelators. Metal chelation during glue secretion, however, may block some of these cross-links. Such robust, non-specific interactions in an aqueous environment are highly unusual for hydrogels and reflect an intriguing cross-linking mechanism.

Cross-linking by protein oxidation in the rapidly setting gel-based glues of slugs

“…When proteins were separated by gel filtration and spotted directly onto a membrane without denaturation, most did not stain for the presence of carbonyls, though there was some staining centered around the fractions containing asmp-40 [which eluted as a giant complex, as seen by Smith et al (Smith et al, 2009)] (Fig.2). There was no detectable staining in the fractions containing asmp-15, despite containing the largest amount of protein.…”

“…In addition, there are likely to be significant amounts of other common divalent ions such as calcium and magnesium, which may also contribute to direct cross-links. Finally, asmp-40 forms large complexes that are insensitive to EDTA (Smith et al, 2009) and do not show a high level of oxidation. These may be cross-linked in a different way.…”

“…Terrestrial slug glue contains significant amounts of zinc, iron, copper and manganese (Werneke et al, 2007). The presence of metals is essential to the glue; metal removal prevents the gel-stiffening activity of the primary crosslinking proteins (Werneke et al, 2007), and disrupts the integrity of the gel (Smith et al, 2009). A major question now is the mechanism by which the metals cross-link the glue.…”

The effectiveness of electrified fencing using copper electrodes for slug (Airon spp.) control with direct electric current and voltage

“…Consistent with this hypothesis, exposure to Cu 2+ was shown to represent a sublethal stressor in pleurocercid snails (Paulson et al, 1983) and was toxic for bivalves and polychaetes as well as gastropods (Kidwai and Ahmed, 1999;Cheung et al, 2002). Smith et al (2009) showed that metal chelation disrupts the crosslinking of macromolecule polymer gels in gastropod mollusks. These adhesive gels carry relatively high percentages of negatively charged amino acids (Pawlicki et al, 2004) that exhibit high-affinity EDTA-sensitive binding of divalent cations such as Ca 2+ and Mg 2+ (Smith et al, 2009) and should therefore be strongly affected by electrolytically produced protons and Cu 2+ .…”

“…Smith et al (2009) showed that metal chelation disrupts the crosslinking of macromolecule polymer gels in gastropod mollusks. These adhesive gels carry relatively high percentages of negatively charged amino acids (Pawlicki et al, 2004) that exhibit high-affinity EDTA-sensitive binding of divalent cations such as Ca 2+ and Mg 2+ (Smith et al, 2009) and should therefore be strongly affected by electrolytically produced protons and Cu 2+ . Accordingly, heavy metals such as Cu 2+ impact gastropod chemoreception, resulting in decreased feeding behavior, foot paralysis and death (Cuthbert et al, 1976).…”