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Biomolecular Materials, Biomineralization and New Materials
Nature builds remarkable high-performance composite materials, such as the exceptionally strong microlaminate of the abalone shell (seen in the pearly background of this figure on the right, and in the electron micrograph at the far left of the same graphic). The precision of this biological nanoscale engineering far exceeds the present capabilities of human engineering. Our interdisciplinary team is discovering the proteins, genes and molecular mechanisms that control the biological nanofabrication of these materials, and using them to develop new routes for synthesis of high-performance composites needed for the technologies of tomorrow. Potential applications include new optoelectronic, microelectronic and catalytic devices (such as the polymetallic crystalline thin-film semiconductor made under the control of proteins purified from the abalone shell, shown in the upper right), and improved biosensors (such as the DNA chip used for robotic diagnostic screening for HIV, other viral and microbial infectious agents and genetically inherited diseases, illustrated in the lower center). [Polymetallic crystalline thin-film courtesy of Dr. Angela Belcher (Acta Mater. 46: 733-736, 1998); DNA chip courtesy of Affymetrix Corp.] Other images are explained below.
Glassy needles of silica made by a marine sponge (above). We recently discovered that protein filaments inside these needles can catalyze the synthesis of opal-like silica and high-performance silicone polymer networks, depending on the substrate provided. Gene cloning revealed the surprising fact that the principal protein (which we named “silicatein”) is highly homologous to a well-known enzyme. These discoveries point to a possible reaction mechanism (confirmed by our recent site-directed mutagenesis studies), and offer the prospect of a new route to the environmentally benign synthesis of high-performance silicon-based materials. (Proc. Natl. Acad. Sci. 95: 6234-6238, 1998; photomicrograph by L. J. Friesen.)
Gene cloning reveals the structures of the proteins controlling biomineralization. Molecular modeling of these proteins (such as the silicatein shown on the left) then guides our site-directed and combinatorial mutagenesis, to produce genetically engineered proteins designed to alter the structures of the composite materials they control.
The abalone shell is a microlaminate composite of calcium carbonate crystals and proteins, with a fracture-toughness 3,000-times greater than that of the crystals alone (right). Although the proteins comprise only a few percent of the mass of the composite, they’re responsible for the tremendous enhancement of strength of the material and the precise control of its unique nanostructure. Using an experimentally tractable “flat pearl” model system we developed for these studies, together with state-of-the-art atomic force microscopy, X-ray diffraction and molecular biology, we’re resolving the proteins, genes and molecular mechanisms responsible for this control. (Nature 371: 49-51, 1994; Proc. Royal Soc. London, B 256: 17-23, 1994; Chem. Matrls. 8: 679-690, 1996; Nature. 381: 56-58, 1996; Acta Mater. 46: 733-736, 1998; Curr. Opin. Colloid Interface Sci. 3: 55-62, 1998)
Growing crystals (each = 0.5 micron thick, forming in conical, tapered stacks) at the growing surface of an abalone pearl (left). The growth of this unusual structure is controlled by three different families of proteins. One of these families forms self-assembled fenestrated sheets that separate the crystal layers and act as molecular stencils, guiding the growth of the crystals from one layer to the next to produce the tapered conical stacks shown here. The stochastic location of nanopores in the fenestrated protein sheets generates the random offset of crystal growth in each successive layer. This offset generates the interdigitation of crystal plates that contributes to the exceptional strength of the composite material. (Chem. Mater. 9: 1731-1740, 1997)
We found that a “genetic switch” controls the abrupt transition from calcite to aragonite synthesis in the abalone shell and flat pearl. Here, proteins purified from the calcite and aragonite crystals are shown to control the “polymorph selection” and atomic lattice orientation of calcite and aragonite crystals produced in vitro, matching with perfect fidelity the control exhibited in vivo. (The rhombohedral crystals were formed under the control of the calcitic proteins; the abrupt transition to the overlying aragonite crystals was produced by addition of the aragonitic proteins.) This observation demonstrated that the “genetic switch” acts by differentially controlling the synthesis of these two different groups of proteins. It also made possible the synthesis of multiphasic microlaminate crystal composites over micron scale dimensions, with perfect control of crystal polymorph and atomic lattice orientation. (Nature. 381: 56-58, 1996) These results then served as the basis for the use of the crystal-controlling proteins to produce polymetallic crystalline thin-films with useful semiconductor and magnetic properties (Acta Mater. 46: 733-736, 1998).
Recent innovations in atomic force microscopy (developed by our close colleague, Professor Paul Hansma, and his students and postdoctorals in Physics) permit us to analyze the molecular-level interactions of the proteins with the atomic surfaces of the growing crystals they control, in real time. (Biophysical Journal 72: 1425-1433, 1997)
Cloning and sequence analysis of its DNA revealed that the “lustrin” proteins from the abalone shell and flat pearl exhibit a unique, alternating modular structure (above). Molecular modeling, analysis of the strength of isolated lustrin molecules with the atomic force microscope, and electron micrographic histochemical localization of the lustrins within the shell all suggest that this modular protein acts as an “elastomeric linker”, holding the crystalline layers together and contributing an elasticity and strength to the microlaminate that were previously unsuspected. (J. Biol. Chem. 272: 32472-32481, 1997)
In our related work with the critters themselves (Click Here) we’ve discovered the signal molecules that induce the larvae of abalones, corals, and several other marine invertebrates to metamorphose. Taking advantage of the fact that this induction synchronously triggers expression of the genes and proteins that commence and control biomineralization, we’ve used this method to clone the DNAs that specifically code for the proteins governing biomineralization. Expression cloning of DNAs coding for the proteins that direct biomineralization in the newly metamorphosed coral is illustrated; the newly formed skeleton, induced at metamorphosis 20 hours previously, is visible in the young coral (and in an electron micrograph ).
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