Sea stars adhere firmly but temporarily to various substrata as a result of underwater efficient adhesive secretions released by their tube feet. Previous studies showed that this material is mainly made up of proteins, which play a key role in its adhesiveness and cohesiveness. Recently, we solubilized the majority of these proteins and obtained 43 de novo-generated peptide sequences by tandem MS. Here, one of these sequences served to recover the full-length sequence of Sea star footprint protein 1 (Sfp1), by RT-PCR and tube foot transcriptome analysis. Sfp1, a large protein of 3,853 aa, is the second most abundant constituent of the secreted adhesive. By using MS and Western blot analyses, we showed that Sfp1 is translated from a single mRNA and then cleaved into four subunits linked together by disulphide bridges in tube foot adhesive cells. The four subunits display specific protein-, carbohydrate-, and metal-binding domains. Immunohistochemistry and immunocytochemistry located Sfp1 in granules stockpiled by one of the two types of adhesive cells responsible for the secretion of the adhesive material. We also demonstrated that Sfp1 makes up the structural scaffold of the adhesive footprint that remains on the substratum after tube foot detachment. Taken together, the results suggest that Sfp1 is a major structural protein involved in footprint cohesion and possibly in adhesive interactions with the tube foot surface. In recombinant form, it could be used for the design of novel sea star-inspired biomaterials.

Intrinsically disordered proteins rich in cationic amino acid groups can undergo Liquid-Liquid Phase Separation (LLPS) in the presence of charge-balancing anionic counterparts. Arginine and Lysine are the two most prevalent cationic amino acids in proteins that undergo LLPS, with arginine-rich proteins observed to undergo LLPS more readily than lysine-rich proteins, a feature commonly attributed to arginine's ability to form stronger cation-π interactions with aromatic groups. Here, we show that arginine's ability to promote LLPS is independent of the presence of aromatic partners, and that arginine-rich peptides, but not lysine-rich peptides, display re-entrant phase behavior at high salt concentrations. We further demonstrate that the hydrophobicity of arginine is the determining factor giving rise to the reentrant phase behavior and tunable viscoelastic properties of the dense LLPS phase. Controlling arginine-induced reentrant LLPS behavior using temperature and salt concentration opens avenues for the bioengineering of stress-triggered biological phenomena and drug delivery systems.

Low molecular weight chitosan (LMW chitosan, ∼5 kDa) potentially has many desirable biomedical applications such as anti-microbial, anti-tumor, and anti-diabetes. Unlike high molecular weight chitosan, LMW chitosan is easily dissolvable in aqueous solutions even at neutral and basic pH, but its dissolution mechanism is not well understood. Here, we measured adhesion and cohesion of molecularly thin LMW chitosan films in aqueous solutions in different buffer pHs (from 3.0 to 8.5) using a surface forces apparatus (SFA). Interestingly, significantly lower adhesion force was measured for LMW chitosan films compared to the high molecular weight chitosan (∼150 kDa) films. Not only the strength of adhesion is lower, but also the critical contact time where adhesion starts to increase with contact time is longer. The results from both the SFA and atomic force microscopy (AFM) indicate that, in physiological and basic conditions, the low cohesion of LMW chitosan due to the stiffness of the chain which cause strong electrostatic correlation energy penalty when they are aggregated. Here, we propose the reduction in cohesion for shorter chitosan (LMW chitosan) as an explanation of its high solubility of LMW chitosan in physiological pHs.

Many marine invertebrates utilize biomacromolecules as building blocks to form their load-bearing tissues. These polymeric tissues are appealing for their unusual physical and mechanical properties, including high hardness and stiffness, toughness and low density. Here, a marine hydroid perisarc of Aglaophenia latirostris was investigated to understand how nature designs a stiff, tough and lightweight sheathing structure. Chitin, protein and a melanin-like pigment, were found to represent 10, 17 and 60 wt.% of the perisarc, respectively. Interestingly, similar to the adhesive and coating of marine mussel byssus, a DOPA (3,4-dihydroxyphenylalanine) containing protein and iron were detected in the perisarc. Resonance Raman microprobe analysis of perisarc indicates the presence of catechol-iron(III) complexes in situ, but it remains to be determined whether the DOPA-iron(III) interaction plays a cohesive role in holding the protein, chitin and melanin networks together.

Chitosan, a load-bearing biomacromolecule found in the exoskeletons of crustaceans and insects, is a promising biopolymer for the replacement of synthetic plastic compounds. Here, surface interactions mediated by chitosan in aqueous solutions, including the effects of pH and contact time, were investigated using a surface forces apparatus (SFA). Chitosan films showed an adhesion to mica for all tested pH ranges (3.0-8.5), achieving a maximum value at pH 3.0 after a contact time of 1 h (Wad ~ 6.4 mJ/m(2)). We also found weak or no cohesion between two opposing chitosan layers on mica in aqueous buffer until the critical contact time for maximum adhesion (chitosan-mica) was reached. Strong cohesion (Wco ~ 8.5 mJ/m(2)) between the films was measured with increasing contact times up to 1 h at pH 3.0, which is equivalent to ~60% of the strongest, previously reported, mussel underwater adhesion. Such time-dependent adhesion properties are most likely related to molecular or molecular group reorientations and interdigitations. At high pH (8.5), the solubility of chitosan changes drastically, causing the chitosan-chitosan (cohesion) interaction to be repulsive at all separation distances and contact times. The strong contact time and pH-dependent chitosan-chitosan cohesion and adhesion properties provide new insight into the development of chitosan-based load-bearing materials.

Complex coacervation driven liquid-liquid phase separation (LLPS) of biopolymers has been attracting attention as a novel phase in living cells. Studies of LLPS in this context are typically of proteins harboring chemical and structural complexity, leaving unclear which properties are fundamental to complex coacervation versus protein-specific. This study focuses on the role of polyethylene glycol (PEG)-a widely used molecular crowder-in LLPS. Significantly, entropy-driven LLPS is recapitulated with charged polymers lacking hydrophobicity and sequence complexity, and its propensity dramatically enhanced by PEG. Experimental and field-theoretic simulation results are consistent with PEG driving LLPS by dehydration of polymers, and show that PEG exerts its effect without partitioning into the dense coacervate phase. It is then up to biology to impose additional variations of functional significance to the LLPS of biological systems.

The byssal threads of the fan shell Atrina pectinata are non-living functional materials intimately associated with living tissue, which provide an intriguing paradigm of bionic interface for robust load-bearing device. An interfacial load-bearing protein (A. pectinata foot protein-1, apfp-1) with L-3,4-dihydroxyphenylalanine (DOPA)-containing and mannose-binding domains has been characterized from Atrina's foot. apfp-1 was localized at the interface between stiff byssus and the soft tissue by immunochemical staining and confocal Raman imaging, implying that apfp-1 is an interfacial linker between the byssus and soft tissue, that is, the DOPA-containing domain interacts with itself and other byssal proteins via Fe3(+)-DOPA complexes, and the mannose-binding domain interacts with the soft tissue and cell membranes. Both DOPA- and sugar-mediated bindings are reversible and robust under wet conditions. This work shows the combination of DOPA and sugar chemistry at asymmetric interfaces is unprecedented and highly relevant to bionic interface design for tissue engineering and bionic devices.

We describe robustly anchored triblock copolymers that adopt loop conformations on surfaces and endow them with unprecedented lubricating and antifouling properties. The triblocks have two end blocks with catechol-anchoring groups and a looping poly(ethylene oxide) (PEO) midblock. The loops mediate strong steric repulsion between two mica surfaces. When sheared at constant speeds of ∼2.5 μm/s, the surfaces exhibit an extremely low friction coefficient of ∼0.002-0.004 without any signs of damage up to pressures of ∼2-3 MPa that are close to most biological bearing systems. Moreover, the polymer loops enhance inhibition of cell adhesion and proliferation compared to polymers in the random coil or brush conformations. These results demonstrate that strongly anchored polymer loops are effective for high lubrication and low cell adhesion and represent a promising candidate for the development of specialized high-performance biomedical coatings.