Design of Polypeptides Self-Assembling into Antifouling Coatings: Exploiting Multivalency

Abstract

We propose to exploit multivalent binding of solid-binding peptides (SBPs) for the physical attachment of antifouling polypeptide brushes on solid surfaces. Using a silica-binding peptide as a model SBP, we find that both tandem-repeated SBPs and SBPs repeated in branched architectures implemented via a multimerization domain work very well to improve the binding strength of polypeptide brushes, as compared to earlier designs with a single SBP. At the same time, for many of the designed sequences, either the solubility or the yield of recombinant production is low. For a single design, with the domain structure B - M - E , both solubility and yield of recombinant production were high. In this design, B is a silica-binding peptide, M is a highly thermostable, de novo-designed trimerization domain, and E is a hydrophilic elastin-like polypeptide. We show that the B - M - E triblock polypeptide rapidly assembles into highly stable polypeptide brushes on silica surfaces, with excellent antifouling properties against high concentrations of serum albumin. Given that SBPs attaching to a wide range of materials have been identified, the B - M - E triblock design provides a template for the development of polypeptides for coating many other materials such as metals or plastics.

Modular Design for Proteins Assembling into Antifouling Coatings: Case of Gold Surfaces

“…The M domain has previously been reported to be highly thermostable . We have also found this to be the case for the M domain in the context of our previously designed silica-coating proteins Figure d shows that, as expected, the same holds for the M domain in the gold-coating proteins: the spectra do not change when heating from 20 to 95 °C and cooling back to 20 °C (20 °C rev).…”

“…To determine the layer thickness of the B - M - E polypeptide brushes self-assembled onto gold surfaces, we used dynamic light scattering (DLS). As shown earlier for silica nanoparticles and the silica-coating B - M - E polypeptides, bridging interactions may lead to particle clustering at very low concentrations, but at higher concentrations, a fully saturated layer develops, leading to a small but measurable increase in the particle hydrodynamic diameter, from which we can estimate the layer thickness.…”

“…According to the Sauerbrey equation, the frequency drop is only linearly related to adsorbed mass in QCM-D for adsorbed layers with low dissipation. Previously, we have found that the silica-binding B - M - E proteins form highly hydrated polypeptide brushes with strong dissipation . Rather than trying to fit our data with complicated models fully accounting for the dissipation, we here simply use the frequency drop as a qualitative measure for adsorbed mass.…”

“…A B - M - E modular design consisting of three domains was proposed, where B is a solid-binding peptide for anchoring to the surface, M is a multimerization domain for increasing the overall protein binding strength to the surface through multivalency, and E is an uncharged hydrophilic elastin-like polypeptide (ELP) , serving as an antifouling block. Compared to first-generation coatings with the simpler B - E design, , the multivalent B - M - E designs assembled into highly stable coatings that could not be displaced, neither by high salt buffers nor by serum albumin protein solutions. Also, the coatings were highly antifouling against high concentrations of serum albumin.…”

“…To replace the original silica-binding domain, we chose a gold-binding peptide that has been extensively studied both by experimental work and by computer simulations, − and which has previously been referred to as GBP1, a non-cysteine peptide with a single-letter amino acid sequence MHGKTQATSGTIQS. Previously, we have used E = (GSGVP) 40 for the antifouling domain. Here, we explore a series of three different lengths of the zwitterionic ELP sequence, E = E Z n = (GKGVP-GDGVP) n /2 with n = 20, 40, or 80.…”