Design of Compact Biomimetic Cellulose Binding Peptides as Carriers for Cellulose Catalytic Degradation

The conversion of biomass into biofuels can reduce the strategic vulnerability of petroleum-based systems and at the same time have a positive effect on global climate issues. Lignocellulose is the cheapest and most abundant source of biomass and consequently has been widely considered as a source for liquid fuel. However, despite ongoing efforts, cellulosic biofuels are still far from commercial realization, one of the major bottlenecks being the hydrolysis of cellulose into simpler sugars. Inspired by the structural and functional modularity of cellulases used by many organisms for the breakdown of cellulose, we propose to mimic the cellulose binding domain (CBD) and the catalytic domain of these proteins by small molecular entities. Multiple copies of these mimics could subsequently be tethered together to enhance hydrolytic activity. In this work, we take the first step toward achieving this goal by applying computational approaches to the design of efficient, cost-effective mimetics of the CBD. The design is based on low molecular weight peptides that are amenable to large-scale production. We provide an optimized design of four short (i.e., ∼18 residues) peptide mimetics based on the three-dimensional structure of a known CBD and demonstrate that some of these peptides bind cellulose as well as or better than the full CBD. The structures of these peptides were studied by circular dichroism and their interactions with cellulose by solid phase NMR. Finally, we present a computational strategy for predicting CBD/peptide–cellulose binding free energies and demonstrate its ability to provide values in good agreement with experimental data. Using this computational model, we have also studied the dissociation pathway of the CBDs/peptides from the surface of cellulose

Changes to the Disordered Phase and Apatite Crystallite Morphology during Mineralization by an Acidic Mineral Binding Peptide from Osteonectin

Noncollagenous proteins regulate the formation of the mineral constituent in hard tissue. The mineral formed contains apatite crystals coated by a functional disordered calcium phosphate phase. Although the crystalline phase of bone mineral was extensively investigated, little is known about the disordered layer’s composition and structure, and less is known regarding the function of noncollagenous proteins in the context of this layer. In the current study, apatite was prepared with an acidic peptide (ON29) derived from the bone/dentin protein osteonectin. The mineral formed comprises needle-shaped hydroxyapatite crystals like in dentin and a stable disordered phase coating the apatitic crystals as shown using X-ray diffraction, transmission electron microscopy, and solid-state NMR techniques. The peptide, embedded between the mineral particles, reduces the overall phosphate content in the mineral formed as inferred from inductively coupled plasma and elemental analysis results. Magnetization transfers between disordered phase species and apatitic phase species are observed for the first time using 2D ¹H–³¹P heteronuclear correlation NMR measurements. The dynamics of phosphate magnetization transfers reveal that ON29 decreases significantly the amount of water molecules in the disordered phase and increases slightly their content at the ordered-disordered interface. The peptide decreases hydroxyl to disordered phosphate transfers within the surface layer but does not influence transfer within the bulk crystalline mineral. Overall, these results indicate that control of crystallite morphology and properties of the inorganic component in hard tissue by biomolecules is more involved than just direct interaction between protein functional groups and mineral crystal faces. Subtler mechanisms such as modulation of the disordered phase composition and structural changes at the ordered–disordered interface may be involved

Interfacial Mineral–Peptide Properties of a Mineral Binding Peptide from Osteonectin and Bone-like Apatite

Osteonectin is a regulator of bone mineralization. It interacts specifically with collagen and apatite through its N-terminal domain, inhibiting crystal growth. In this work, we investigated the interface formed between the mineral and an acidic peptide, ON29, derived from the protein’s apatite binding domain. The structural properties of the peptide bound to the mineral and the mineral–peptide interface are characterized using NMR and computational methods. A biomaterial complex is formed by precipitation of the mineral in the presence of the acidic peptide. The peptide gets embedded between mineral particles, which comprise a disordered hydrated coat covering apatite-like crystals. ³¹P SEDRA measurements show that the peptide does not affect the overall proximity between phosphate ions in the mineral. {¹⁵N}¹³C REDOR measurements reveal an α-turn in the center of the free peptide, which is unchanged when it is bound to the mineral. {³¹P}¹³C REDOR and ¹H–¹³C HETCOR measurements show that Glu/Asp carboxylates and Thr/Ala/Val side chains from ON29 are proximate to phosphate and hydroxyl groups in the mineral phases. Predictions of ON29’s fold on and off hydroxyapatite crystal faces using ROSETTA-surface are used to model the molecular conformation of the peptide and its apatite-binding interface. The models constructed without bias from experimental results are consistent with NMR measurements and map out extensively the residues forming an interface with apatitic crystals