Polymorph crystal selection by n16, an intrinsically disordered nacre framework protein

Framework proteins are a subclass of mollusk shell nacre-associated polypeptides that form supramolecular assemblies with β-chitin and other matrix proteins. These macromolecular assemblies manage the energetics of aragonite polymorph nucleation, and thus, there is keen interest in understanding the molecular characteristics of framework proteins. Here, we report the mineralization activity, oligomerization, and structural features of a recombinant framework nacre protein n16, isoform 3 (r-n16.3, Japanese pearl oyster Pinctada fucata). We find that r-n16.3 assembles in mineralization solutions to form spheroidal-fibril and mineralized thin film assemblies, in addition to spherical vaterite mineral deposits and aragonite single crystal deposits that possess unusual texture and layered morphologies. The oligomerization of r-n16.3 is spontaneous over the pH range 5-8.5, and protein particle sizes are observed to increase in radii when Ca(II) is present. Bioinformatics studies reveal that the r-n16.3 molecule is intrinsically disordered (random coil) and possesses residual α helix and β sheet structure. Experimentally, we confirmed that the secondary structure of apo-r-n16.3 assemblies is largely disordered (50% random coil, 20% β strand, 8% α helix). However, in the presence of high Ca(II) concentrations, we observe IDP disorder-to-order transformations that increase β turn structure and decrease random coil, α helix, and β strand contents. We conclude that r-n16.3 is an intrinsically disordered oligomeric nacre framework protein that nucleates vaterite and single crystal aragonite in vitro and possesses target-specific IDP disorder-to-order transformation capabilities in response to Ca(II).