C. Steven Sikes

CARBONIC ANHYDRASE AND CARBON FIXATION IN COCCOLITHOPHORIDS1

Rates of carbon fixation in coccolithophorids in culture, unlike many other algae, are carbon limited at ambient levels of dissolved inorganic carbon (DIC). Apparently, plants often rely on activity of carbonic anhydrase (CA) to raise the level of CO2 in cells and achieve carbon saturation. However, CA activities in the coccolithophorids, Coccolithus (= Emiliania) huxleyi Lohmann and Hymenomonas (=Cricosphaera) carterae Braarud, were either not detectable or very low compared to activities in other systems, including other algae, higher plants, and representative animals. Furthermore, additions of CA to medium with 2 mM DIC at pH 8.1 resulted in nearly 30% enhancement of photosynthesis, but not coccolith formation. Although carbon fixation in coccolithophorids can be suppressed by the CA inhibitor acetazolamide, studies of CaCO3 nucleation revealed a non‐specific effect of the inhibitor. Using a 30 min assay based on pH decreases accompanying loss of dissolved. CO32‐, inhibition of crystal formation in the absence of CA at 1 mM acetazolamide was demonstrated for decalcified crab carapace, a tissue with which normal CaCo3 deposition in vitro has been shown. The results suggest only a minor role for CA in coccolithophorids.

Evaluation of calcium binding by molluscan shell organic matrix and its relevance to biomineralization

Abstract 1. 1. EDTA used to dissolve mollusc shells associates with soluble matrix (SM) material and persists in SM preparations despite exhaustive conventional dialysis. 2. 2. EDTA-free oyster or clam SM material binds little calcium at physiological ionic strength. 3. 3. It is concluded that residual EDTA or failure to determine binding in physiological media may lead to binding artefacts and that hypotheses which rely on SM calcium binding to explain initiation of biomineral formation need to be re-evaluated. 4. 4. It is hypothesized that crystal binding, rather than calcium binding, by SM is responsible for initiation of biomineral formation.

Adsorption and modification of calcium salt crystal growth by anionic peptides and spermine

SummarySynthetic polyanions, including peptide analogs of naturally occurring proteins, have been shown to inhibit the nucleation and growth of calcium salt crystals. The binding characteristics of polyaspartate and aspartate-serine copolymers to calcium carbonate (calcite) and hydroxyapatite (HAP) are presented here. The binding is related to dosedependent inhibition of crystal growth measured by constant composition assays. Peptide phosphorylation had little effect on binding affinity or crystal growth inhibition with either calcium salt. Spermine was able to reduce hydroxyapatite crystal growth but with lower efficacy than the polyanionic peptides. Spermine reversed some of the HAP growth inhibition produced by an anionic peptide. Binding of a labeled polyanion was reduced by a similar anionic peptide at all concentrations of the label, however, spermine reduced binding only at higher concentrations of the labeled polyanion. The data support the presence of multiple binding site classes on HAP surfaces, some inaccessible to polycations and some at which both polyanions and polycations can bind.

Atomic-scale imaging of albite feldspar, calcium carbonate, rectorite, and bentonite using atomic-force microscopy

Atomic force microscopy (AFM) was used to investigate the (010) surface of Amelia albite, the basal and (001) planes of CaCO3 (calcite), and the basal planes of rectorite and bentonite. Atomic scale images of the albite surface show six sided, interconnected en-echelon rings. Fourier transforms of the surface scans reveal two primary nearest neighbor distances of 4.7 and 4.9 +/- 0.5 angstroms. Analysis of the images using a 6 angstroms thick projection of the bulk structure was performed. Close agreement between the projection and the images suggests the surface is very close to an ideal termination of the bulk structure. Images of the calcite basal plane show a hexagonal array of Ca atoms measured to within +/- 0.3 angstroms of the 4.99 angstroms predicted by x-ray diffraction data. Putative images of the (001) plane of carbonate ions, with hexagonal 5 angstroms spacing, are also presented and discussed. Basal plane images of rectorite show hexagonal symmetry with 9.1 +/- 2.5 angstroms spacing, while bentonite results reveal a 4.9 +/- 0.5 angstroms nearest neighbor spacing.

Atomic force microscopy and molecular modeling of protein bound to calcite surfaces

Proteins that regulate inorganic crystal formation in organisms are in some instances irreversibly bound to the mineral phase. They are clearly resolved by atomic force microscopy, easily withstanding the forces of the AFM probe in contact mode. This is demonstrated in AFM images of proteins attached to fragments of calcite oyster shell. The proteins may take specific positions on the crystal surface, in effect becoming part of the surficial lattice, stereochemically matching the functional groups of the protein with lattice positions of the constituent ions of the crystal. Requirements for adsorbate binding that is strong enough for AFM viewing in contact mode are discussed, including comparisons of ionically driven binding of oyster shell protein to calcite and hydrogen-bonded antifreeze proteins to ice.