ABSTRACT
A previous study used atomic force microscopy saw-tooth retraction curves to characterize the adhesive mucilage pads of the diatom Toxarium undulatum. The major mucilage component consisted of adhesive nanofibers (ANFs) made up of modular proteins arranged into cohesive units, each containing a set number of modular proteins aligned in parallel. This study shows that T. undulatum adhesive mucilage is a biocomposite containing four additional adhesive components, including single modular proteins that are likely to be the structural units from which the ANFs are assembled. Two further distinct supramolecular assemblies were observed to coexist with ANFs (ANFs II and III), along with a continuum of single modular proteins through oligomers made up of varying numbers of modular proteins arranged in parallel. All components of the adhesive biocomposite produce a characteristic force spectrum with the same interpeak distance (35.3 +/- 0.3 (mean +/- SE) nm), suggesting they are derived from discrete supramolecular assemblies of the same modular protein, but they are distinguishable from one another based on the rupture force, persistence length, and interpeak force measured from their saw-tooth curves.
Subject(s)
Diatoms/metabolism , Proteins/chemistry , Cells, Cultured , Diatoms/cytology , Microscopy, Atomic Force , Proteins/metabolism , Tissue AdhesionsABSTRACT
The adhesive and mechanical properties of a cell-substratum adhesive secreted by live diatom cells were examined in situ using atomic force microscopy. The resulting force curves have a regular saw-tooth pattern, the characteristic fingerprint of modular proteins, and when bridged between tip and surface can repeatedly be stretched and relaxed resulting in precisely overlaying saw-tooth curves (up to approximately 600 successive cycles). The average rupture force of the peaks is 0.794 +/- 0.007 (mean +/- SE) nN at a loading rate of 0.8 microm/s and the average persistence length is 0.026 +/- <0.001 (mean +/- SE) nm (fit using the worm-like chain model). We propose that we are pulling on single adhesive nanofibers, each a cohesive unit composed of a set number of modular proteins aligned in register. Furthermore, we can observe and differentiate when up to three adhesive nanofibers are pulled based upon multimodal distributions of force and persistence length. The high force required for bond rupture, high extensibility (approximately 1.2 microm), and the accurate and rapid refolding upon relaxation, together provide strong and flexible properties ideally suited for the cell-substratum adhesion of this fouling diatom and allow us to understand the mechanism responsible for the strength of adhesion.
Subject(s)
Algal Proteins/chemistry , Algal Proteins/ultrastructure , Cell Adhesion Molecules/chemistry , Cell Adhesion Molecules/ultrastructure , Diatoms/enzymology , Nanostructures/chemistry , Nanostructures/ultrastructure , Adhesiveness , Cells, Cultured , Computer Simulation , Elasticity , Enzyme Activation , Models, Chemical , Models, Molecular , Particle Size , Peptide Mapping , Protein Conformation , Tensile StrengthABSTRACT
Polysaccharides from the red alga Phacelocarpus peperocarpos were extracted with hot water, clarified, and precipitated with 2-propanol. The native preparation was highly sulfated (36.2% w/w). Alkali modification decreased the sulfate content by 2.0% w/w. The alkali-modified polysaccharide is composed mostly of galactose (Gal. 51 mol%) and 3,6-anhydrogalactose (AnGal, 41 mol%), with minor amounts of a mono-O-methylgalactose (MeGal, 1 mol%), xylose (Xyl, 6 mol%), and glucose (Glc, 1 mol%). The FTIR spectrum of the alkali-modified polysaccharide resembled kappa-carrageenan with absorption at 930 cm-1 (indicative of AnGal) and 850 cm-1 (Gal 4-sulfate). However, an additional, major band of absorption occurred sulfate ester substitution at O-6 of at 820 cm-1, indicating the presence of equatorial sulfate ester substitution at O-6 of Gal residues. A combination of linkage and 13C NMR spectroscopic analyses showed that the polysaccharide was composed predominantly of a novel repeating-unit, O-beta-D-galactopyranosyl 4,6-disulfate)-(1-->4)-3,6-anhydro-alpha-D-galactopyranose. Minor structural variations also occurred, including alternative patterns of sulfation and the presence of terminal Xylp. The location of the terminal Xylp residues was not certain but evidence supported their attachment at O-3 of some 4-linked Galp residues. The cell-wall galactans remain unchanged during the life cycle of the alga.
