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Review

Spider silks and their applications


Jonathan A. Kluge, Olena Rabotyagova, Gary G. Leisk and David L. Kaplan
Departments of Biomedical Engineering, Chemistry and Mechanical Engineering, Tufts University, Medford, MA 02155, USA

Spider silks are characterized by remarkable diversity in their chemistry, structure and functions, ranging from orb web construction to adhesives and cocoons. These unique materials have prompted efforts to explore potential applications of spider silk equivalent to those of silkworm silks, which have undergone 5000 years of domestication and have a variety of uses, from textiles to biomedical materials. Recent progress in genetic engineering of spider silks and the development of new chimeric spider silks with enhanced functions and specic characteristics have advanced spider silk technologies. Further progress in yields of expressed spider-silk proteins, in the control of self-assembly processes and in the selective exploration of material applications is anticipated in the future. The unique features of spider silks, the progress and challenges in the cloning and expression of these silks, environmentally triggered silk assembly and disassembly and the formation of bers, lms and novel chimeric composite materials from genetically engineered spider silks will be reviewed. Introduction Silk has captured the imagination of humankind for millenia, largely due to the unrivaled visual and functional properties of silk ber and the unique structures that have been generated by various silk-producing species in nature. These structures include amazing orb web structures spun to capture prey, cocoons to house developing offspring, adhesives used to anchor webs and brous tethers to capture ying prey. Despite signicant interest in other silk sources, such as spider silk, silkworms have been the traditional source exploited. One reason that spider silk has lagged behind is that silkworms are fairly easy to domesticate, whereas spiders cannot be housed in high densities (Encyclopdia Britannica, retrieved December 27th, 2007, http://www.britannica.com/eb/ article-6677). In addition, whereas one silkworm cocoon yields 600 to 900 m of ber, only 137 m of ber can be reeled from the ampullate gland of a spider and only 12 m of silk is found in a complete spider web [1]. Finally, many types of silk are utilized in web construction from a single spider. Therefore, the best option for moving the application of spider silks forward is the pursuit of biotechnological means of generating source material. Recent advances in genetic engineering have led to increased insight into silk proteins and the structural organization of spider-silk-encoding genes. With this knowledge, heterologous expression of spider-silk proteins in a range of host systems has been achieved, as well as the formation of new materials from recombinant-DNA-derived spider-silk
Corresponding author: Kaplan, D.L. (David.Kaplan@tufts.edu).

proteins. These advances might make spider silk a viable choice in many new application areas, heretofore the domain of silkworm silk. Chemistry, structure and function The molecular structure of silk consists of regions of protein crystals separated by less organized protein chains. The primary structural modules give rise to diverse secondary structures that, in their turn, direct functions of different silks. As the most heavily studied secondary structure of silks, crystalline b-sheets contribute to the high tensile strength of silk bers. Beta sheets form through natural physical crosslinking of amino acid sequences, which in spider and silkworm silk consist of multiple repeats of mainly alanine, glycine-alanine, or glycine-alanine-serine. The non-crystalline regions of silk are commonly made up of: (i) b-spirals similar to a b-turn composed of GPGXX repeats (where X is mostly glutamine) and (ii) helical structures composed of GGX [2,3]. These semi-amorphous regions provide silk with elasticity. For example, the agelliform silk from Nephila clavipes is rich in the GPGXX motif, and this sequence results in a highly elastic ber that functions in prey capture. In addition to the crystalline and semi-amorphous regions, non-repetitive regions are present at the amino- and carboxyl termini of the proteins. Although the impact of these termini on mechanical response is not fully understood, it has been speculated that they might play a role in the controlled assembly of silk proteins [4,5]. Each silk-producing creature synthesizes silk proteins that offer a rich diversity of primary sequences and secondary structures. For example, the common amino acid modules in the silk bers synthesized by the Araneomorphae (true spiders) can be grouped into four categories: poly-Ala, poly-Ala-Gly, GPGXX, GGX and a spacer sequence [6]. Most recently, Garb and colleagues characterized six novel silk proteins from the Mygalomorphae (tarantulas) that do not contain these four categories found in true spider spidroins. These newly-characterized tarantula silks, as a result, do not possess high tensile strength and elasticity. This nding supports the hypothesized role of poly-Ala and GPGXX modules in forming dragline and ageliform silks [7]. In most of the above cases, the fundamental process of silk protein self-assembly into functional materials remains consistent, with the more hydrophobic domains, mainly the alanine, glycine-alanine and glycine-alanineserine repeats, driving the process. In most spider silks, bsheet formation is achieved in a spinning duct caused by the progressive loss of water in the gland and alignment of the hydrophobic regions during ow (Figure 1) [812]. Exceptions are the more hydrophilic silks, such as those

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0167-7799/$ see front matter 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.tibtech.2008.02.006 Available online 25 March 2008

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Figure 1. Structural hierarchy in silk assembly related to assembly into fibers. (a) (i) Spider-silk proteins consist of repeats of amino acid sequences that self-assemble into b-sheets. This self assembly is driven by hydrogen bonding and also by hydrophobic regions. These interactions result in the formation of inter- or intra-molecular protein chain interactions. The b-sheet structures further assemble into soft micelles in a manner that excludes the hydrophilic ends to the perimeter. The interiors of the micelles contain water (blue regions in the figure) due to the presence of small spacers that are more hydrophilic than the dominant hydrophobic domains. This does not represent a multi-shell structure; rather, it is caused by the partitioning of the hydrophilic chain ends to the surface of the micelles and the location to the interior of the large and dominant hydrophobic domains and small hydrophilic spacers. With increasing protein concentration, micelles transform into gel-like states leading to metastable liquid crystalline structures. (ii) Triggers, such as physical shearing, or environmental factors, such as low pH, methanol, ultrasonication and electric fields, convert the gel states and liquid crystals into a more stable b-sheet structure. The resulting fibrils emerging from spinning ducts are combined into higher ordered structures as naturally constructed webs or cocoons. (b) Molecular structure of a spider-silk protein. In silks that are used for web architecture and other strong fibers, the underlying molecular structure consists mainly of hydrophobic regions (illustrated by thin black lines). These large hydrophobic regions are interspersed with small hydrophilic spacers (illustrated by gray vertical rectangles) and are flanked by nonrepetitive domains at the N- and C-termini (green horizontal rectangles).

involved in adhesion, wherein charge interactions can play a more dominant role than the hydrophobic interactions. The self-assembly without chemical crosslinking provides stability while still allowing enzymatic digestion [4] or slow degradation under appropriate environmental conditions [13]. Sources and cloning of spider silks The spider silks that have been most extensively studied to date with recombinant DNA technology include those from N. clavipes (Major Spidroin dragline types I and II MaSpI and MaSpII), Araneus diadematus (dragline ADF-3 and ADF-4) and N. clavipes agelliform [1422]. Two main approaches have been used to obtain the gene encoding spider silks: (i) isolation of native spider silk amino acid sequences from peptide digests, followed by back-translation into the corresponding DNA sequence and chemical synthesis of oligonucleotides to represent the sequence and (ii) generation of cDNAs encoding the spider silk from mRNAs that were isolated directly from silk-producing glands. Because spider silk sequences are formed from repeated polypeptides, chemically synthesized oligonucleotides encoding these polypeptide units can be used as building blocks for ligation into multiple repeats to generate longer silk-encoding genes. The advantage of

using synthetic oligonucleotides is that codons can be optimized for different expression systems in an attempt to improve nal protein yield. The rst approach (i) was recently successfully employed in obtaining the full length spider silk gene sequence and anking regions from the black widow spider (Latrodectus hesperus) [23,24]. The elucidation of the genetic organization of spider silk genes has led to new insights into evolutionary biology based on their complex organization of introns and exons [6,25 29]. These insights point to the importance of these remarkable gene structures in fostering stability and high levels of expression in the native hosts. However, this very structural design that has served well in generating the remarkable mechanical properties of silks remains an impediment to the large-scale expression of spider silks in simpler host systems, such as Escherichia coli. This impediment to scaling up has prompted efforts to explore a range of different host systems, including yeast (e.g. Pichia pastoris), insect cells (e.g. sf9), bovine mammary epithelial alveolar cells, baby hamster kidney cells, transgenic plants such as Arabidopsis, soybean, potato and tobacco, transgenic mammals such as mouse and goat and, nally, transgenic silkworms [8,14,15,17,19,21,22,3033]. An important result was the generation of transgenic mice that carried spider dragline silk genes under control of the whey acidic protein (WAP)
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promoter [30]. These mice were able to produce ADF-3 and MaSpI silk proteins and secreted these in their milk over generations. However, despite the use of the various host systems described above, the molecular sizes of the expressed spider-silk proteins are much smaller compared to that of native proteins due to issues of gene stability and the repetitive nature of the genetic sequences involved. To date, a successful expression of full length spider silk clones has not been achieved. To fully recapitulate silk properties, all protein domains present in the native proteins are thought to be crucial, and the absence of some protein domains therefore severely affects the quality of the resulting silk, either because of improper assembly or loss of material properties. Commercial applications of recombinant spider silks therefore remain limited because of the inability to produce sufcient quantities of silk proteins at a reasonable cost and with an accurate molecular weight. Because production costs for plant expression are estimated to be a fraction of those for bacterial fermentation [21], transgenic plants might prove a feasible option once the purication issue has been solved. Processing spider silks into biomaterials One attractive application of spider silks is to emulate the diverse material functions of this family of proteins as a source of novel biomaterial designs. Insight into the assembly and processing of spider-silk proteins into various material forms has been a longstanding focus and has allowed the broadening of the eld of applications for silks in general. Specically, medical devices and tissue

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engineering applications are perhaps the most promising areas for the utilization of spider silks. Recent progress with reprocessed or native silkworm silk bers has been realized, and similar approaches could be used with spider silks when they become available in sufcient quantities. For example, in ligament tissue engineering, a combination of ber twisting and braiding of silkworm silk bers was able to direct stem-cell- and ligament-cell-based reconstruction through the alignment and mechanical strength of the twisted silk structure [34,35]. Recently, spider silk bers were manually collected from the major ampullate dragline and seeded with human Schwann cells to demonstrate biocompatibility, suggesting a promising strategy for future treatment of peripheral nerve injuries [36]. Various studies have been conducted to assess the solubility and solution structure of genetically engineered spider silks, including variants of ADF-3 and ADF-4 that adopted a random coil structure similar to analogs of major ampullate gland proteins in N. clavipes [37]. Different secondary structure results were obtained for protein analogs of MaSpI [16]. To control solubility of spider-silk proteins, genetically engineered variants have been generated with environmentally regulated molecular triggers based on either chemical or biochemical reactions (Figure 2) [3841]. These systems were based on N. clavipes dragline silk analogs, one with the insertion of methionine residues to serve as redox triggers and the other based on the inclusion of an enzymatic phosphorylation site for cyclic AMP-dependent protein kinase phosphorylation. Both systems were reversible (oxidation/reduction;

Figure 2. Molecular triggers for environmental control of b-sheet assembly. To control assembly of spider silks, different modes of genetic modification have been employed that allow a controllable assembly or disassembly of silk b-sheets. (a) The consensus repeat of spider dragline silk from N. clavipes is shown. b-sheet forming regions are indicated by the brick wall icon, which denotes their insoluble state when assembled. (b) A phosphorylation site was introduced by the addition of two new codons into the sequence to provide a suitable recognition site for kinase reaction. This modification generated a controllable b-sheet forming region, based on the presence of the kinase. The phosporylation prevented assembly of the silk protein, whereas dephosphorylation by a phosphatase promoted the assembly of the protein into b-sheets. (c) Introduction of methionine encoding sequences near the b-sheet-forming region generated an oxidation/reduction trigger for the formation of a b-sheet region. Upon chemical oxidation of the methionine side chain, b-sheet formation was prevented, while chemical reduction of the side chain promoted b-sheet formation.

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phosphorylation/dephosphorylation). The use of molecular triggers provides options to regulate the assembly of spider silks for both fundamental biophysical studies and for potential biomaterial applications. Formation of novel biomaterials from recombinant spider-silk proteins Silk proteins have been shown to solubilize in water, organic solvents and ionic liquids, indicating the versatile options available, and they can then be processed into new biomaterials, including bers, lms, gels, porous sponges and other related systems (Figure 3) [37,42]. The resulting structures and functions of the obtained materials are directly controlled by the content and distribution of crystalline b-sheets, a process that can be controlled by the mode of processing. A variety of environmental factors, including solvent, pH, water, concentration of protein and salt, inuence the processing and assembly of spider silk in vivo and in vitro [40]. Attempts to process spider-silk proteins into distinct classes of functional materials have been underway since the rst successful expression systems were realized. Highlighted below are advances in the eld that were based primarily on the processing of genetically engineered proteins derived from spider silk, with insight from silkworm silks serving as a starting point. Fibers and textiles The combination of high strength, toughness and light weight makes spider silks attractive for high-performance ber and composite applications [43] and for biomedical applications. Whether the silk material is to be used as an individual ber or woven into a textile structure, it is critical that production techniques are developed to

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generate long lengths of material in sufcient quantity and with mechanical performance that is at least equal to the native spider silks. Traditionally, silkworm silks have been the focus of research to generate silk ber materials. Techniques to form bers from silkworm proteins, such as solvent extrusion, electrospinning and microuidic approaches, might be appropriate for spider-silk proteins as well (Figure 4). The advantages and limitations of each system might determine their use in specic applications or their commercial exploitation. Silkworm silk solutions can be solubilized in aqueous solvents, including LiBr [44] and N-methylenemorpholineN-oxide (NMMO) [45], and subsequently formed into bers by a wet solvent-based process [46,47]. However, relatively poor ber mechanical properties and brittle behavior were found when compared to native bers (400 MPa compared to upwards of 700 MPa reported for native B. mori silks). Recent efforts using spider silk, however, have shown promise. Spider silk bers have been formed through the electrospinning of spin dopes prepared by dialyzing genetically engineered silk (MaSpI protein analogs) in urea containing Tris buffer and salts. This aqueous method has generated bers that are 1060 mm in diameter and that exhibit molecular orientation and mechanical properties similar to natural spider silk [43]. Electrospinning of spin dopes derived from ADF-3 recombinant spider-silk protein has generated ber diameters up to 40 mm. Although ber toughness and modulus compared favorably to native A. diadematus dragline silk, tenacity was lower [14]. Electrospinning was also used to generate bers with an average diameter of 300 nm from N. clavipes MaSpI silk dope prepared in HFIP (hexauoroisopropanol). It was shown that control of conformation can be

Figure 3. Processing of silk into new biomaterials. Silk fibers, such as native spider silks, silkworm silks and recombinant silk proteins, can be solubilized with a variety of solvents, including formic acid, HFIP, calcium nitrate, lithium salts or ionic liquids. Once solubilized, the silk protein solutions can be processed into the range of different structures shown. Portions of the figure were adapted from [35] and are reprinted with permission of Elsevier Limited.

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Figure 4. Schematic representation of current silk fiber-forming techniques. (a) Solvent extrusion is performed by drawing the fiber through a coagulation bath in a controlled manner. Microfluidic devices use a contracting channel and multiple solvent inputs to regulate the geometry and chemistry of the resulting fiber. Electrospinning processes combine strong voltage gradients and syringe pump extrusion and result in either random or aligned fiber deposition. (b) Characteristics of the different fiberforming processes. To date, electrospinning has proved to be the most useful approach, mainly because of the low amounts of spider silk materials needed and the utility of the resulting electrospun fiber mats for cell- and tissue-culture studies. Portions of the figure were adapted from [46] and are reprinted with permission of John Wiley & Sons, Inc.

achieved and that molecular alignment might be possible, potentially offering improved ber properties [48]. Films Films have been cast using HFIP solutions that contain recombinant forms of the dragline spider-silk proteins ADF-3 and ADF-4. The transparent lms, ranging in thickness from 0.5 to 1.5 mm, could be made water-insoluble through the use of potassium phosphate or methanol, which converted the proteins secondary structure from ahelix to b-sheet. When this treatment was applied to the recombinant form of the ADF-4 protein, a level of chemical stability in certain denaturants was achieved that rivaled the stability of native dragline silk. Cloning strategies and the ability to modify the lm surfaces for attachment of functional molecules make these materials promising for applications such as wound dressings and enzyme immobilization scaffolds [49,50]. Hydrogels and porous sponges Hydrogels are utilized in the eld of tissue engineering to form porous but stable tissue scaffolds. By adding methanol to recombinant forms of the dragline silk protein ADF4, the silk has been shown to self-assemble into nanobers with diameters of 3 nm and lengths less than 1 mm. Over the course of a few days, these nanobers transformed into a hydrogel ber network. These hydrogels exhibited a non248

linear viscoelastic material response with low stiffness and strength. Cross-linking was induced by visible light after applying ammonium peroxodisulfate and tris(2,20 -bipyridyl)dichlororuthenium (II) to the surface of the hydrogel. Cross-linked hydrogels demonstrated fairly linear material response with much greater modulus and strength response. Given the superior mechanical response and the stability of the hydrogel over weeks, these materials are suitable for tissue engineering applications [51]. Sponge-like, porous three-dimensional structures are also important in tissue engineering. Such structures act as a scaffold to support cells, allow transport of nutrients and metabolic wastes and promote tissue development [52,53]. While development of silkwormsilk-based sponges has progressed for such applications, the use of spider silk has lagged behind. In one application of spider silk, spider cocoon silk was used to create porous scaffolds for tissue engineered cartilage. Through a process of solubilizing of the cocoon silk in lithium bromide (LiBr), mixing with salts and methanol treatment, porous scaffolds were generated that sustained chondrocyte cell growth, which could lead to articular cartilage development [54]. Microcapsules Microcapsules of the recombinant form of ADF-4 spider silks were developed by controlling silk self-assembly at an

Review
oilliquid interface. The resulting b-sheet-rich thin polymer shells had high mechanical stability, with wall thicknesses on the order of 50 nm and diameters between 1 and 30 mm. In addition, the properties of the microcapsules, including constrained degradation response to tissue-specic enzymes, were easily controlled. These materials, therefore, offer promise for a range of applications, from drug delivery to microreactor design [55]. Functionalization of spider silk novel composites Surface functionalization is an important strategy for modifying the exterior of biomaterials to inuence cell and tissue development, and this strategy has been applied to silk surface chemistry [42]. Surface modication usually targets carboxylic acid groups on the amino acids in the protein. Silkworm broin silk has been modied by coupling enzymes such as horseradish peroxidase, cell binding domains such as Arg-Gly-Asp (RGD) peptides and cell signaling factors such as parathyroid hormone (PTH) and bone morphogenetic protein-2 (BMP-2) [44,56,57] to improve cell interactions and function or to successfully form gradients on the silk material surface. However, in silkworm silk broin only 3.3% of the amino acids have carboxyl side groups, whereas this number is signicantly higher in spider silks, which offer the advantage of forming higher degrees of substitution with

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cell-modifying functional groups. For example, MaSp1 has 11.7% of aspartic acid and glutamic acid residues, which are theoretically available for chemical modications [58]. So far, the dragline silks ADF-3 and ADF-4 from the garden spider Araneus diadematus have been functionalized by coupling lms formed from the proteins with uorescein and b-galactosidase using carbodiimide chemistry via EDC (1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide) [49]. This approach offers high levels of surface decoration with enzymes, which could be used for biosensors and reactive media and to inuence cell and tissue functions. Genetically engineered spider silks can offer the added benets of providing designed functionalities encoded during gene construction. Strategies employed for spider silks have included the incorporation of cell binding domains, such as RGD [59], or domains that interact selectively with inorganic components to generate novel organicinorganic composite material systems [60,61], where N. clavipes provides the spider silk component. Highly tailored hybrid or chimeric spider silks containing cell integrin binding domains (RGD) [59] and domains that were able to selectively undergo mineralization with silica [60] and hydroxyapatite [61] were generated (Figure 5). The composite morphology and structure could be manipulated by controlling processing conditions to produce lms and bers. The control over the size of the

Figure 5. Generation of chimeric silk proteins as a tool to expand their functional features. (a) The consensus spider silk sequence that constitutes the core repeats of chimera is shown. (b) Examples of protein regions used for the generation of chimeric silk proteins. The R5 component of silaffin (sequence shown) was fused to spider dragline silk and was able to foster the polymerization of silica precursors to form glassified materials. The C-terminal domains from dentin matrix protein 1 (DMP 1), a protein found in the mineralized tissue of teeth and responsible for the nucleation and growth of hydroxyapatite, could also be fused to the spider silk and was shown to nucleate and form hydroxyapatite-containing spider silk in film form. The third example shows the addition of a cell binding domain, RGD from fibronectin, which is responsible for cell binding via membrane integrin receptors. This resulted in cell adhesion on silk films. The last example shows the formation of block copolymers formed from silkworm silk and elastin for biomaterials used as drug delivery systems. (c) The basic cloning procedure involving synthetic oligonucleotides is illustrated; this procedure leads to the purifcation of silk proteins derived from recombinant DNA that have new functions. These novel proteins can be used to generate silk materials with improved functionalities, such as improved cell binding features, glassified silk fibers with increased stiffness and hydroxyapatite mineralized biomaterials potentially useful for bone-related repairs. Portions of the figure were adapted from [60] and [61] and are reprinted with permission of Elsevier Limited and Proc. Natl. Acad. Sci. U. S. A., respectively. The spider picture was taken by Josh Hillman and Emily Earp (http://FLNature.org/).

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inorganic domains, as well as the molecular-level interactions between the organic and inorganic domains in these composites, offers a biomimetic approach toward tougher and stiffer silk composite materials that might be potentially useful in hard tissue remodelling or in adhesive llers. These results suggest that chimeric spider-silk proteins might provide new uses for spider silk in biomedical and other specialty materials. Further benets of this approach might arise due to the ability to process and assemble these novel composite materials in aqueous ambient processing conditions. Conclusions The path ahead for spider-silk proteins remains challenging, mainly due to the inadequate supply of full length proteins that are required for a full exploration of the most interesting applications. Thus, currently, the more readily available silkworm silk is most widely applied in the biomedical area. Over the last decade there has been considerable progress in understanding the gene organization of spider silk. The cloning and expression techniques for spider silks have been improved, and the self-assembly and processing of spider silk into many material formats is now better understood. However, further achievements are required, such as the expression of full length spider silk genes to fully embrace the complex sequence structure important for accurate assembly and function of spider silks, as well as the establishment of efcient protein purication protocols, especially from transgenic plants. It will also be important to further understand and exploit the processing of these proteins in aqueous environments to fully recapitulate the native process used by spiders to organize these proteins into functional materials. This is essential because b-sheet formation dictates the resulting material properties, and their features are inuenced directly by the presence of water and the rate of water removal during formation of the protein into biomaterials. An important benet of using spider silks is their range of material characteristics, which arise from the extensive array of spider-silk proteins that are used throughout the normal life-cycle of these organisms. This is a feature that distinguishes spider silk from B. mori silkworm silk, which contains only one single type of silk. The inuence of genetic variation, and thus protein chemistry, on the material properties and applications for different spider silks needs to be further investigated. One path to achieving this improved understanding is via the use of synthetic genes to encode the key sequences in the various silks for their assembly into functional materials. Full length clones or synthetic analogs that encompass all key sequence components in a spider silk will be required for these studies. Furthermore, these full length clones will facilitate improved aqueous processing and the assembly into different materials originating from these proteins. Achieving this goal will open the door to the use of these diverse spider-silk protein systems in a range of novel applications, from glues to tougher composite materials and lightweight webbing. At present, novel hybrid or chimeric spider silk systems have been developed that are feasible for a variety of biomedical processes, such as the use of silk-inorganic composites to
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regulate cellmaterial interfacial interactions, to control bonding to tissues and to modulate degradation proles. These applications are expected to be realized in the near future because only relatively low quantities of silk proteins are needed. However, applications of spider silks in commodity materials, such as textiles, high performance composite materials and durable and tough materials in general, will require the success of robust and low cost expression systems. The development of a range of recombinant spider silks, possibly generated in a combinatorial mode, would offer unique insights into a variety of processes, such as protein preparation and protein polymer processing, with potential for improved biomaterial designs. Furthermore, it remains to be seen whether silkworm silk variants could also be produced to fully exploit the range of designs that have evolved in nature, such as orb webs, cocoons and glues. The ability to mimic these features through genetic engineering and protein processing offers possibilities for expanding the variety of biologically derived materials that can be applied for medical and non-medical uses. Interest in silk has increased with a near explosion of biomaterials research and applications, and other natural silk sources, such as honeybees, hornets and solitary wasps, have also attracted attention. For example, it has been shown that honeybee silk is composed of coiled-coil proteins. This results in characteristics that are different from b-sheet silks, such as differences in processing and assembly options and in the resulting material properties (e.g. increased toughness) [62]. Further exploitation of the aqueous processing and assembly of highly hydrophobic silk proteins into material structures can also be considered as a path into green chemistry in a broader context. Because the silk proteins, as highly hydrophobic polymers, can be processed, assembled and formed into robust materials using only water, this sets out a path for the exploration of other hydrophobic polymers in a similar fashion, even if they are of purely synthetic origin. Thus, spider silk has the potential to have an impact not only on materials science and engineering but also on green chemistry and biomedicine after anticipated scientic and engineering progress is achieved.
Acknowledgements
We thank our many collaborators, students and colleagues who have been instrumental in the studies described in part here. Grant support on silkrelated systems is greatly appreciated and has included, at various times, the National Institutes of Health, the National Science Foundation and the Air Force Ofce of Scientic Research. Special thanks go to Andy Lovinger and Hugh Delong for their foresight and support.

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Trends in Biotechnology

Vol.26 No.5

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