Incorporation of a Recombinant Biomineralization Fusion Protein into the Crystalline Lattice of Calcite

Abstract

High-resolution synchrotron X-ray powder diffraction (XRD) combined with the Rietveld refinement method and confocal laser scanning microscopy (CLSM) were utilized in this study to elucidate the interaction between a recombinant biomineralization protein (perlucin) fused to green fluorescent protein (GFP) and synthetic calcite. Although recombinant perlucin is insoluble, its solubility was increased via fusion to the highly soluble GFP. We demonstrate that GFP-perlucin derivatives become incorporated into the calcite structure and induce concentration-dependent anisotropic lattice distortions along the host’s c-axis. In contrast, GFP alone is hardly incorporated at all. The observed lattice distortions and peculiar microstructure of the crystals are comparable to those previously observed in biogenic calcite. Taking advantage of biotechnology to optimize individual protein properties, such as the solubility of an otherwise insoluble protein derivative, is a promising route toward the synthesis of new and improved biocomposite materials.

Structure and Properties of Nanocomposites Formed by the Occlusion of Block Copolymer Worms and Vesicles Within Calcite Crystals

“…[ 25,32 ] Nevertheless, comparison of the calcite prisms from a range of mollusks has revealed species-specifi c variations in the microstrain and macrostrain, [ 25,33 ] where these have been attributed to differences in the compositions of the macromolecules, rather than the amounts occluded. [ 34 ] Investigations have therefore often focused on calcite crystals coprecipitated with extracted molecules, where XRD spectra from calcite crystals occluding the small protein perlucin [ 24 ] showed smaller peak shifts than calcite precipitated in the presence of "caspartin," a 17 kDa protein extracted from the calcitic prisms of Pinna nobilis . [ 32 ] Purely synthetic systems-such as the copolymer worms and vesicles studied here-offer an alternative and effective strategy for determining the relationship between the chemical structure, size, and shape of additives and the structure and properties of nanocomposite crystals.…”

“…Peak broadening is commonly reported on heating both synthetic and biogenic calcite crystals containing occluded organic phases, where this is attributed to the creation of new interfaces. [ 9,24,25 ] The vesicle/calcite crystals were therefore subjected to isochronous annealing for 30 min at intervals of 100 °C from 100-600 °C and patterns were recorded after the samples had cooled to room temperature after each heating cycle. Peak shifts to smaller d -values began on heating to 300 °C, where this continues on heating to higher temperatures ( Figure 5 a,b).…”

3D visualization of additive occlusion and tunable full-spectrum fluorescence in calcite

“…Our study provides new insight into the occlusion of small organic molecules (here fluorescent dyes) within calcite single crystals, demonstrating the existence of zoning and the relationship between occlusion and changes in crystal morphologies. It is noted that proteins tagged with fluorescent dyes have been occluded within polycrystalline and single-crystal calcite, but zoning was not observed45. The dye Congo Red has also been reported to occlude within calcite46, and a recent AFM study has showed that this dye stabilizes the polar step edges along the [010] direction of calcite (10.4)47.…”

Wenn fluoreszierende Proteine und Weißlicht emittierende Dioden aufeinandertreffen

“…Dadurch wird die Anwendung von FPs in Bauteilen wie OLEDs eingeschränkt. [89,91] Insbesondere die FP-Immobilisierung wurde sowohl in organischen als auch in anorganischen Stoffen wie a-Lactose-Monohydrat, [92,93] Polymermatrizen [24] und Calcit realisiert. In dieser Hinsicht sind wässrige Lçsungen von FPs und/oder Trockenfilme nicht die beste Wahl fürd ie Einbindung in Beleuchtungskçrper oder elektronische Bauelemente,s ondern es müssen andere Matrizen verwendet werden, die die natürliche Struktur der FPs über längere Zeiträume aufrechterhalten.…”

“…[87] Zum anderen ist die Verwendung von FPs in Beleuchtungsanwendungen nur sinnvoll, wenn ihre Struktur und ihre spektralen Eigenschaften über lange Zeit stabil gehalten werden kçnnen, um idealerweise ein Bauteil mit langer Lebensdauer zu realisieren. [91] Die Photostabilität, insbesondere hinsichtlich der Fluoreszenzwellenlänge (Farbton) und der PLQY (Intensität), ist ein weiterer überaus wichtiger Aspekt, der bei der Einbindung von FPs als aktive Bestandteile in Beleuchtungskçrpern berücksichtigt werden muss.D iesbezüglich gibt es verschiedene photodynamische Prozesse,d ie diese Eigenschaften beeinflussen kçnnen. Bei der Auswahl der Matrix fürdie Immobilisierung der Proteine im Allgemeinen, und der FPs im Besonderen, müssen die Affinitätf ürd as Protein, die Verfügbarkeit von reaktiven funktionellen Gruppen, die mechanische Stabilität, die Festigkeit und die Biokompatibilitäti nB etracht gezogen werden.…”