The Intrinsically Disordered C-RING Biomineralization Protein, AP7, Creates Protein Phases That Introduce Nanopatterning and Nanoporosities into Mineral Crystals

Abstract

We report an interesting process whereby the formation of nanoparticle assemblies on and nanoporosities within calcite crystals is directed by an intrinsically disordered C-RING mollusk shell nacre protein, AP7. Under mineralization conditions, AP7 forms protein phases that direct the nucleation of ordered calcite nanoparticles via a repetitive protein phase deposition process onto calcite crystals. These organized nanoparticles are separated by gaps or spaces that become incorporated into the forming bulk crystal as nanoporosities. This is an unusual example of organized nanoparticle biosynthesis and mineral modification directed by a C-RING protein phase.

Intrinsically disordered proteins and biomineralization

“…Calcium carbonate formation in shells, for example, is regulated by IDPs. One shell protein, AP7, first shown to form supramolecular-assemblies and nucleate single crystals of aragonite [36] is also involved in the regulation of calcite growth [37]. The C-RING-like sequence is an important site for AP7 self-association and mineral nucleation; self-assembling, to form protein phases that congregate amorphous calcium carbonate nanoparticles, into hybrid, protein–mineral supramolecular networks, and create nano-porosities on the surfaces of the calcite that forms.…”

“…The C-RING-like sequence is an important site for AP7 self-association and mineral nucleation; self-assembling, to form protein phases that congregate amorphous calcium carbonate nanoparticles, into hybrid, protein–mineral supramolecular networks, and create nano-porosities on the surfaces of the calcite that forms. Chang et al [37] speculated that the repeated deposition of AP7 on the calcite surface allows for the creation of these nano-porosities. The ability of IDPs to interact with and regulate more than one mineral phase is another important theme that will occur throughout this discussion.…”

“…A recent neutron spectroscopy, small-angle X-ray and neutron scattering of osteopontin-calcium phosphate interactions, confirmed that the 1-149 fragment of osteopontin maintained its flexible linear chain structure in solution, stabilizing ACP particles [115]. Additionally, stabilization of calcium-carbonate nanoparticles and creation of nano-porosities within them is due to the IDP mollusk protein, AP7 [37], discussed above, implying this could be a common IDP –mineral regulation mechanism. As reviewed elsewhere [116] the initial mineral that forms, be it an amorphous liquid, or pre-nucleation chains or clusters, may exhibit polyamorphism.…”

Synergistic Biomineralization Phenomena Created by a Combinatorial Nacre Protein Model System

“…12-19 These families include the framework, intracrystalline, and pearl/reparative proteomes 12-26 that are localized throughout the nacre layer. Recent experiments have shown that 4 representative nacre proteins (AP7, 13-15 PFMG1, 18 n16.3, 16,17 Pif97 12 ) exist as intrinsically disordered, aggregation-prone species that spontaneously form hydrogels or protein phases under both low and high ionic strength conditions. 12-17 These phases have been shown to organize and assemble amorphous calcium carbonate (ACC) nanoparticles during the early stages of calcium carbonate nucleation and control the formation kinetics of pre-nucleation clusters (PNCs), the predecessors of ACC.…”

“…For this, we created a model combinatorial system consisting of two intrinsically disordered nacre proteins, the intracrystalline shell nacre protein, AP7 ( Haliotis rufescens , 66 AA, C-RING domain 31-66, chemically synthesized form = cAP7) 13-15 and the pearl-associated nacre protein, PFMG1 ( Pinctada fucata , 116 AA, pseudo-EF hand domain 61-110, bacterial recombinant form = rPFMG1)(Figure S1, Supporting Information). 18,21 The C-RING-containing cAP7 forms hydrogels that induces the formation of intracrystalline nanoporosities and nanotexturing or nanopatterning on the exposed surfaces of calcite.…”