Organic biopolymers of venus clams: Collagen-related matrix in the bivalve shells with crossed-lamellar ultrastructure.

BACKGROUND: Biochemical studies and spectroscopic techniques have shown that chitin-silk fibroins are common in nacroprismatic bivalve shells. However, the nature of organic biopolymers in the less well studied shell architectures, such as crossed lamellar shells, remain unknown. Here, two venus shells, Callista disrupta and Callista kingii, with crossed lamellar ultrastructure have been studied. METHODS: We employed thermal gravimetric analysis, optical-, confocal- and scanning electron-microscopes, gel-sodium dodecyl sulfate (gel-SDS), FTIR, ultra-performance liquid chromatography and high-performance anion-exchange chromatography system with pulsed amperometric detection to analyse organic macromolecules in the shells. RESULTS: Thermal analysis showed a low concentration of organic macromolecules in C. disrupta (1.38 wt%) and in C. kingii (1.71 wt%). A combination of biochemical protocols, including Calcofluor White staining and FTIR spectroscopic assessment, indicate that amino-polysaccharide chitin together with proteins, are present in the organic scaffolding of the shells. Scanning electron microscope of insoluble acid biopolymer extracts as well as FTIR technique show that the hierarchical structural organizations of organic biopolymers consist collagen-related matrix. Our histochemical fixing and staining techniques reveal many discrete proteins and glycoproteins from soluble organic macromolecules on the gel-SDS. We show here 'singlet' and 'doublet' glycosaminoglycan bands that are far above 260 kDa. GENERAL SIGNIFICANCE/CONCLUSIONS: The presence of collagen matrix in Callista shells shows promise for the new source of biomaterials. Most importantly, the structural organization of the proteinaceous motif is predominantly helical structures and not silk-fibroin unlike in nacreous bivalve shells.

Characterization of organophosphatic brachiopod shells: spectroscopic assessment of collagen matrix and biomineral components.

The shells of linguloid brachiopods such as Lingula and Discinisca are inorganic-organic nanocomposites with a mineral phase of calcium phosphate (Ca-phosphate). Collagen, the main extracellular matrix in Ca-phosphatic vertebrate skeletons, has not previously been clearly resolved at the molecular level in organophosphatic brachiopods. Here, modern and recently-alive linguliform brachiopod shells of Lingula and Discinisca have been studied by microRaman spectroscopy, Fourier transform infrared spectroscopy, field emission gun scanning electron microscopy, and thermal gravimetric analysis. For the first time, biomineralized collagen matrix and Ca-phosphate components were simultaneously identified, showing that the collagen matrix is an important moiety in organophosphatic brachiopod shells, in addition to prevalent chitin. Stabilized nanosized apatitic biominerals (up to ∼50 nm) permeate the framework of organic fibrils. There is a ∼2.5-fold higher wt% of carbonate (CO3 2-) in Lingula versus Discinisca shells. Both microRaman spectroscopy and infrared spectra show transient amorphous Ca-phosphate and octacalcium phosphate components. For the first time, trivalent moieties at ∼1660 cm-1 and divalent moieties at ∼1690 cm-1 in the amide I spectral region were identified. These are related to collagen cross-links that are abundant in mineralized tissues, and could be important features in the biostructural and mechanical properties of Ca-phosphate shell biominerals. This work provides a critical new understanding of organophosphatic brachiopod shells, which are some of the earliest examples of biomineralization in still-living animals that appeared in the Cambrian radiation.