Pif80 is a key protein for the formation and organization of mineral precursor in pearl oyster nacre biomineralization.

Nacre is the inner layer of the mollusc shell and provides exceptional toughness via its highly organized organic−inorganic composite structure. Pif is an organic matrix protein from the nacreous layer of the pearl oyster Pinctada fucata and exhibits regulatory behavior in nacre formation. Here, we investigated features of Pif97, the Nterminal of Pif, using a recombinant form of Pif97 produced in Escherichia coli. We observed that recombinant Pif97 was able to efficiently form a complex with calcium ions. Additionally, recombinant Pif97 showed both in vitro growth inhibition of thermodynamically stable calcite, stabilization of amorphous calcium carbonate, and exclusive binding affinity to metastable aragonite and chitin. These results imply the participation of Pif97 in the calcification of nacre including the association of the inorganic phase and polysaccharide template. We propose that recombinant Pif97 has inherent characteristics of the native form, which are significant for interrelating with organic matrix and inorganic calcium carbonate during nacre biomineralization.

The development of carbonic anhydrase (CA)-based materials for the environment-friendly sequestration of carbon dioxide (CO 2 ) under mild conditions would be highly valuable for controlling emissions to the environment and for producing value-added chemicals. Here, a highly tough and stable CA-encapsulating silk protein hydrogel was developed as a robust biocatalyst for CO 2 sequestration through a bioinspired dual-crosslinking strategy that employed photoinduced dityrosine chemical crosslinking followed by dehydration-mediated physical crosslinking. The target enzyme was efficiently encapsulated in the silk hydrogel with 60% retention of the activity of free CA, and the encapsulated CA exhibited excellent overall multi-use, storage and thermal stabilities. The dual-crosslinked CA-encapsulating silk hydrogel exhibited a significant compressive modulus, which surpassed the moduli of most traditional and double-network hydrogels as well as those of enzyme-encapsulated hydrogels. This hydrogel also showed high resiliency and elasticity and outstanding structural stability. Importantly, the dual-crosslinked CA-encapsulating silk hydrogel facilitated the sequestration of CO 2 into calcium carbonate with high CO 2 hydration activity. Thus, the unique combination of bioinspired dual-crosslinking with silk fibroin protein and CA enzyme demonstrates the successful application of this protein hydrogel as a promising biocatalyst for CO 2 sequestration by showing high activity, strong mechanical properties and outstanding structural stability.Lyophilized dc-ngCA-silk hydrogels were immersed in 20 mM Tris-HCl (pH 8.3) for 1 day. The weights of the swollen silk hydrogels were measured Biocatalytic tough protein hydrogel CS Kim et al

BackgroundUnique adhesive and biocompatibility properties of mussel adhesive proteins (MAPs) are known for their great potential in many tissue engineering and biomedical applications. Previously, it was successfully demonstrated that redesigned hybrid type MAP, fp-151, mass-produced in Gram-negative bacterium Escherichia coli, could be utilized as a promising adhesive biomaterial. However, purification of recombinant fp-151 has been unsatisfactory due to its adhesive nature and polarity which make separation of contaminants (especially, lipopolysaccharide, a toxic Gram-negative cell membrane component) very difficult.ResultsIn the present work, we devised a high resolution purification approach to secure safety standards of recombinant fp-151 for the successful use in in vivo applications. Undesirable impurities were remarkably eliminated as going through sequential steps including treatment with multivalent ion and chelating agent for cell membrane washing, mechanical cell disruption, non-ionic surfactant treatment for isolated inclusion body washing, acid extraction of washed inclusion body, and ion exchange chromatography purification of acid extracted sample. Through various analyses, such as high performance liquid chromatographic purity assay, limulus amoebocyte lysate endotoxin assay, and in vitro mouse macrophage cell tests on inflammation, viability, cytotoxicity, and apoptosis, we confirmed the biological safety of bacterial-derived purified recombinant fp-151.ConclusionsThrough this purification design, recombinant fp-151 achieved 99.90% protein purity and 99.91% endotoxin reduction that nearly no inflammation response was observed in in vitro experiments. Thus, the highly purified recombinant MAP would be successfully used as a safety-secured in vivo bioadhesive for tissue engineering and biomedical applications.

A thin film was formed through in vitro CaCO3 crystallization in the presence of complex coacervates, which was expected to be planar and poorly crystalline CaCO3 guided at the interface of two immiscible liquid phases upon complex coacervation.