Editorial pubs.acs.org/journal/abseba
Editorial: Special Issue on Designer Protein Biomaterials
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materials exhibit reversible, thermoplastic behavior that allows them to be easily molded or extruded at modest temperatures. Moreover, SRTs also fully dissolve in weak acid buffers into their suckerin components, which can then be reconstituted into nativelike materials by solvent casting. Many of the interesting properties of the native SRT material can also be recapitulated using recombinant suckerins, providing opportunities for tailored materials. The serendipitous discovery of this protein material, discussed at length in the review, highlights once again the importance of basic science (in this case marine biology) on recent advances in the biomaterials field. Burkhard and co-workers have pioneered the development of self-assembling protein nanoparticles (SAPNs) for drug delivery and vaccine applications. These SAPNs are composed of de novo designer peptides comprised of two amphiphilic αhelical domains connected by a short linker sequence into a Vshaped construct. On their own, the two α-helical domains were designed to form distinct pentameric and trimeric coiledcoil motifs. This allows the SAPN peptides to self-assemble into capsid-like icosahedra that preserve the 3- and 5-fold symmetry of their α-helical domains. Such capsid-like peptide assemblies have a central core that may be used for encapsulation of therapeutic agents and can also easily be modified to include functional sequences displayed on the nanoparticle periphery for targeted delivery or immunological signaling. In the current article,3 a comprehensive in silico and in vitro combinatorial design study of sequence selection is presented for formation of stable protein nanoparticles below the cutoff size (∼15 nm) required to minimize unwanted immunogenic response. The power of in silico screening of a large number of potential sequence variants for desired biophysical properties is a notable feature of this article and points to a clear trend in the use of computational methods for the design of biomaterials with specific biophysical and functional attributes in the future. Biologically synthesized designer proteins often are challenging to produce and purify in the large quantities required for biomedical applications. In the next article, Joshi and coworkers4 describe a simple and robust filtration method to directly purify protein nanofibers produced by a bacterial host. The protein studied was the amyloid-forming CsgA expressed in modified E. coli strains in the form of curli amyloid protein fibers. As these fibers are relatively stable in the presence of chaotropic solutions (such as GdmCl) and surfactant solutions, the authors were able to develop a processing protocol to obtain purified mats of protein nanofibers directly from bacterial suspensions, using sequential filtering from solutions with added guanidinium chloride, nucleases, and sodium dodecyl sulfate. Moreover, this scheme was successfully extended to chimeras of CsgA and the SpyCatcher domain, demonstrating a route to custom functionalized protein
here is a long history of protein-based biomaterials, starting from proteins purified from natural sources (collagen, elastin, and silk are prominent examples). The rise of genetic engineering and molecular biotechnology not only made it possible to allow precise control over the sequence and chain length of such proteins derived from nature but also enabled the design and engineering of artificial proteins for the construction of protein biomaterials, opening the door to designer protein biomaterials. Over the last two decades, significant progress has been made in designer protein biomaterials, and the ability to design and create a vast number of different proteins with desired properties for potential use in biomaterials applications is firmly established, whereas the main challenges in the field perhaps have more to do with issues limiting the successful application of these materials. Recent developments in this direction have been facilitated by technological developments (large scale computation resources for in silico protein design, knowledge-based tools provided by genomics and proteomics, essential materials processing technologies such microfabrication and bioprinting) and also by gains in fundamental understanding that are occurring through the interaction of the biomaterials community with researchers in other fields of engineering, medicine, and science. This special issue has a selection of articles and reviews on designer protein biomaterials that highlight some of these recent developments in the field. Advances in fundamental, molecular-level understanding of the structure and function of animal tissues have provided fresh ideas for new biomimetic systems. Elastomeric proteins are an important class of proteins found in nature that have inspired numerous biomimetic protein polymers for biomaterials applications. In the first contribution to this special issue, Muiznieks and Keeley1 provide a comprehensive review of the origins and properties of natural elastomeric systems (including elastins, resilins, abductins, collagens, and silks), and of engineered biomimetic materials based on these natural proteins. The focus of this review is on elucidating the general relationships between the sequence and structure of the proteins and the mechanical properties of the materials they form, and using this knowledge in the design of new biomimetic elastomeric proteins. This is a compelling paradigm for biomaterials development. The power of a multidisciplinary approach, involving bioinformatics, computational biophysics, molecular biology and zoology, also clearly emerges from this review. The next contribution is a review by Hiew and Miserez2 on a fascinating natural protein-based biomaterial found in the teethlike protrusions of the suckers lining the appendages of squid and cuttlefish. These so-called sucker ring teeth (SRT) are tough, high modulus elastic materials with remarkable properties. They are almost entirely composed of a family of proteins called suckerins, which have some sequence similarity to silks. Indeed, in the SRT the suckerins have a high fraction of β sheet domains connected by amorphous, disordered regions, as in native silk fibers. However, unlike silk fibers these © 2017 American Chemical Society
Special Issue: Designer Protein Biomaterials Received: April 24, 2017 Published: May 8, 2017 658
DOI: 10.1021/acsbiomaterials.7b00256 ACS Biomater. Sci. Eng. 2017, 3, 658−660
ACS Biomaterials Science & Engineering
Editorial
are required, for instance to allow for cell motility and proliferation within the matrix. The article by Kiick and coworkers7 describes a clever use of arrested liquid−liquid phase separation as a means to produce microstructured composite hydrogels of resilin-like polypeptides (RLP) and polyethylene glycol (PEG). In this scheme, the hydrogels are formed by cross-linking the primary amines of RLP and amine-terminated PEG with tris(hydroxymethyl phosphine) during phase separation of an initially homogeneous mixture. By varying the concentrations of RLP and PEG in the 2-phase region of the mixture, they were able to create hydrogels with a tunable distribution of RLP-rich domain sizes. As the viscoelastic properties of the RLP and PEG materials are substantially different, this leads to materials with locally varying mechanical properties, a highly desirable attribute in some soft tissue regeneration applications. The approach presented in this paper is clearly transferable to numerous protein-based biomaterial systems (including more complex multicomponent systems) and has potential for in situ cell encapsulation with suitable cross-linking chemistries. Surface modification of nonbiological materials (e.g., synthetic polymers, ceramics, and metals) using proteins or peptides is an established approach for controlling biological responses to otherwise incompatible or bioneutral materials. In this area, designer protein biomaterials play an increasingly important role. The article by Scheibel and co-workers8 describes the utility of recombinant spider silk proteins, based on a consensus sequence in the core domain of the dragline silk of the European garden spider A. diadematus, for enhancing the biocompatibility (through controlled cell adhesion) of three conventional catheter materials (polyurethane, polytetrafluoroethylene, and silicone). A simple, robust layer-by-layer dipcoating approach is described that utilizes alternating layers of anionic and cationic silk mimetic proteins. The resulting coatings were shown to be mechanically robust, stable in culture media, and moderately resistant to protease activity. Moreover, in the case of anionic silk top layers, the coated catheter materials were shown to inhibit the adhesion of a variety of cells, and thereby reducing foreign body responses such as periprostetic capsular fibrosis. Notably, such coatings can also be designed to encourage cell adhesion, either nonspecifically by using cationic variants or specifically by insertion of selected cell binding motifs in the silk sequence, indicating that this is broadly useful approach to surface functionalization. The final contributions to this special issue are two comprehensive and complementary reviews of the use of natural and biomimetic proteins in regenerative medicine applications. The first review by Miranda-Nieves and Chaikof9 discusses recent developments in the use of collagen and elastin biomaterials for tissue engineering. Through a systematic recapitulation of collagen and elastin biosynthesis and the biophysical properties of these systems, a case is made for the physiological importance of composite materials of collagen and elastin. The authors subsequently provide an overview of fabrication methods for processing these protein biomaterials to produce hybrid cell-matrix constructs for tissue engineering, and present selected case studies for the use of these systems in regeneration of cardiovascular, liver, musculoskeletal, and skin tissues. The second review by Weiss and co-workers10 provides a detailed progress report on the use of both natural and synthetic biomaterials for the development of small-diameter vascular grafts. Following a thorough discussion of the synthetic
nanofiber materials. Although this work is specific for the amyloid-forming CsgA protein system, it should be broadly useful for any sufficiently stable fibrillar protein assembly. The potential sequence complexity of protein and peptide systems allows the design of dynamic biomaterials whose properties can show dynamic changes in response to environmental stimulus. Well-studied examples include biomaterials that assemble/disassemble in response to changes of solution conditions such as pH, temperature, or ionic strength. Of particular recent interest are more complex responses to solution constituents. For example, the article by Li and coworkers5 describes protein hydrogels that utilize metal chelation to regulate mechanical properties via protein folding/unfolding events. This was achieved by engineering an artificial protein folding switch, whose two conformations (folded versus unfolded) can be regulated by the chelation of Ni2+. The folding and unfolding of the artificial protein folding switch leads to an effective change of the chain length between two cross-linking points, and consequently the cross-linking density of the protein hydrogel is modified. Via this mechanism, the mechanical properties of the resultant protein hydrogels can be dynamically regulated by divalent metal ions. This scheme provides a reversible way for in situ regulation of the mechanical properties of protein hydrogels, and may provide a convenient approach to design dynamic protein hydrogels for biological applications. Another interesting class of responsive biomaterials involves self-assembly in response to a second essential component, such as another complementary protein, polymer or particle, as a route to forming a responsive composite material. The article by Heilshorn and co-workers6 describes the use of designed molecular recognition between peptide-functionalized hydroxy apatite nanoparticles and target recombinant proteins to stimulate the assembly of hybrid protein-nanpoparticle hydrogels. This development represents a biofunctional generalization of the established polymer−nanoparticle (PNP) paradigm for self-healing materials, in which the peptidefunctionalized nanoparticles serve as transient, multivalent cross-links between protein chains. As with traditional PNP systems, such a hydrogel can be temporarily fluidized by the application of shear stress, because of the noncovalent nature of the peptide−protein interactions, facilitating the use of the system as an injectable biomaterial. However, unlike traditional PNP systems, the peptide−protein interactions are specific in nature, allowing for precise control of the assembly process. Moreover, through the addition of other recombinant proteins (or peptides) with competitive binding sites, the cross-link density in the protein-nanoparticle hydrogel can be modulated at will. The utility of this nanocomposite hydrogel system for regenerative medicine applications was demonstrated by its dual use in bone regeneration as (i) a thixotropic medium to encapsulate and deliver adipose-derived stem cells (ASCs) into a biodegradable porous scaffold and (ii) an in situ, osteoconductive matrix (because of the hydroxy apatite nanoparticles) to guide stem cell differentiation. However, the general scheme will likely prove to be useful for many applications requiring an injectable, biofunctional cell encapsulation material. Although the biochemical, local structural and mechanical properties of protein-based biomaterials are highly controlled by the choice of protein sequence, it is more challenging to encode desired mesoscale morphology into the protein sequence. In many applications, highly porous biomaterials 659
DOI: 10.1021/acsbiomaterials.7b00256 ACS Biomater. Sci. Eng. 2017, 3, 658−660
ACS Biomaterials Science & Engineering
Editorial
and biopolymer systems in current use or development for vascular grafts, the authors argue that the ideal materials for these applications are likely to be composites of synthetic polymer materials that possess optimized mechanical properties and extracellular matrix proteins (elastin/collagen hybrids) that provide biocompatibility and integration with native cellular components of the vasculature. The utility of hybrid synthetic/ biological materials is a common theme in regenerative medicine applications. Developers of future protein biomaterial systems would do well to make integration with both biological tissues and synthetic materials a design priority.
James L. Harden Department of Physics, University of Ottawa
Hongbin Li
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Department of Chemistry, University of British Columbia
AUTHOR INFORMATION
ORCID
James L. Harden: 0000-0002-4118-7209 Hongbin Li: 0000-0001-7813-1332 Notes
Views expressed in this editorial are those of the authors and not necessarily the views of the ACS.
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REFERENCES
(1) Muiznieks, L. D.; Keeley, F. W. Biomechanical Design of Elastic Protein Biomaterials: A Balance of Protein Structure and Conformational Disorder. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/ acsbiomaterials.6b00469. (2) Hiew, S. H.; Miserez, A. Squid Sucker Ring Teeth: Multiscale Structure−Property Relationships, Sequencing, and Protein Engineering of a Thermoplastic Biopolymer. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00284. (3) Dey, R.; Xia, Y.; Nieh, M.-P.; Burkhard, P. Molecular Design of a Minimal Peptide Nanoparticle. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00243. (4) Dorval Courchesne, N.-M.; Duraj-Thatte, A.; Tay, P-K. R.; Nguyen, P. Q.; Joshi, N. S. Scalable Production of Genetically Engineered Nanofibrous Macroscopic Materials via Filtration. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00437. (5) Kong, N.; Fu, L.; Peng, Q.; Li, H. Metal Chelation Dynamically Regulates the Mechanical Properties of Engineered Protein Hydrogels. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00374. (6) Parisi-Amon, A.; Lo, D. D.; Montoro, D. T.; Dewi, R. E.; Longaker, M. T.; Heilshorn, S. C. Protein−Nanoparticle Hydrogels That Self-assemble in Response to Peptide-Based Molecular Recognition. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00286. (7) Lau, H. K.; Li, L.; Jurusik, A. K.; Sabanayagam, C. R.; Kiick, K. L. Aqueous Liquid−Liquid Phase Separation of Resilin-Like Polypeptide/Polyethylene Glycol Solutions for the Formation of Microstructured Hydrogels. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/ acsbiomaterials.6b00076. (8) Borkner, C. B.; Wohlrab, S.; Möller, E.; Lang, G.; Scheibel, T. Surface Modification of Polymeric Biomaterials Using Recombinant Spider Silk Proteins. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/ acsbiomaterials.6b00306. (9) Miranda-Nieves, D.; Chaikof, E. L. Collagen and Elastin Biomaterials for the Fabrication of Engineered Living Tissues. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00250. (10) Hiob, M. A.; She, S.; Muiznieks, L. D.; Weiss, A. S. Biomaterials and Modifications in the Development of Small-Diameter Vascular Grafts. ACS Biomater. Sci. Eng. 2017, DOI: 10.1021/acsbiomaterials.6b00220.
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DOI: 10.1021/acsbiomaterials.7b00256 ACS Biomater. Sci. Eng. 2017, 3, 658−660
