Sequence Directed Crystal Design in the Prismatic Layer of the Mollusk

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INTELLECTUAL MERIT: Mollusk shells consist of two nanostructured, opposing layers. The inner, or pearl-like nacreous, layer consists of calcium carbonate in the aragonite crystal morphology and confers fracture toughness on the composite. The outer, or prismatic, layer consists of long prismatic columns of single crystal calcium carbonate in the calcite modification that confers puncture and crack propagation resistance. Both the nacreous and prismatic layers arise initially during shell development from a precursor phase, consisting of amorphous calcium carbonate. Here it is proposed to investigate the detailed mechanism by which the prismatic layer is assembled through the intermediacy of specific negatively charged aspartic acid and glutamic acid-rich proteins, known collectively as Asprich, thought to direct the mineralization process. Proteins of this class have highly conserved sequences at the N and C termini. A peptide (Fragment 1) from the N-terminal domain promotes radial rhombohedral crystal arrangements, while Fragment 2 from the C-terminus promotes porosities or voids at crystal surfaces. This project will utilize synthetic peptides representing Fragments 1 and 2 to understand how conserved sequences control crystal growth and design. In particular, the correlation of primary sequence organization, amino acid identity, secondary structure, and metal ion binding capacity with crystal morphology selection and growth kinetics will be investigated. Scanning electron and atomic force microscopies will be used to examine crystal morphologies. Various biophysical techniques (CD, NMR, isothermal titration calorimetry, mass spectrometry, nano-SIMS, X-ray diffraction, computer simulation) will provide means to examine the properties of the Asprich domains with regard to Ca(II) and Mg(II) binding thermodynamics, 2- and 3D structure, surface electrostatics, and crystal binding and occlusion within the calcite matrix. Collaborations with Brookhaven National Laboratory (E. DiMasi) and Lawrence Livermore National Laboratory (C. Orme) will provide access to equipment not available locally to the project.BROADER IMPACTS: The project provides a platform for highly multidisciplinary training of graduate students ranging from peptide synthesis, to various microscopies, to biophysical characterization with synchrotron X-ray diffraction to NMR, nano-SIMS, and isothermal titration calorimetry. The project has the potential to advance the understanding of biomineralization and to provide models for biomimetic composite fabrication. The PI is a participant in the New York Academy of Sciences High School Science Summer Camp and invites 1-2 high school students to the lab to do research experiments. He also participates in the Partners in Science high school science teacher research program which introduces NYC high school science teachers to research projects during their summer vacation.