Spider silk, an ancient biomaterial, has many qualities worth replicating. With the use of geneti... more Spider silk, an ancient biomaterial, has many qualities worth replicating. With the use of genetic modification, relatively large amounts of spider silk protein have been produced utilizing transgenic goat milk. Access to this protein has allowed us to create spider silk films and hydrogels. Applications for these products are far reaching, from football helmets to drug release capsules. Using mammalian cells, it has been proven that spider silk is not only biocompatible, but that it degrades at an ideal rate. As medical advances have taken place, the ability to create scaffolds for biomedical research and tissue implantation have increased. Spider silk films and hydrogels are an ideal material to consider for such scaffolds. This project focuses on discovering and improving mammalian cell growth on spider silk films and hydrogels so as to create optimal scaffolds. We plan to see beneficial cell growth and attachment by virtue of the composition of our spider silk films and hydrogels compared to our controls
Spider silk, which has remarkable mechanical properties, is a natural protein fiber produced by s... more Spider silk, which has remarkable mechanical properties, is a natural protein fiber produced by spiders. Spiders cannot be farmed because of their cannibalistic and territorial nature. Hence, large amounts of spider silk cannot be produced from spiders. Genetic engineering is an alternative approach to produce large quantities of spider silk. Our group has produced synthetic spider silk proteins in E.coli to study structure/function and to produce biomaterials comparable to the silks produced by orb-weaving spiders. Here we give a detailed description of our cloning, expression, and purification methods of synthetic spider silk proteins ranging from ∼30 to ∼200 kDa. We have cloned the relevant genes of the spider Nephila clavipes and introduced them into bacteria to produce synthetic spider silk proteins using small and large-scale bioreactors. We have optimized the fermentation process, and we have developed protein purification methods as well. The purified proteins are spun into fibers and are used to make alternative materials like films and adhesives with various possible commercial applications.
Spider silks are well known for their extraordinary mechanical properties. This characteristic is... more Spider silks are well known for their extraordinary mechanical properties. This characteristic is a result of the interplay of composition, structure and self-assembly of spider silk proteins (spidroins). Advances in synthetic biology have enabled the design and production of spidroins with the aim of biomimicking the structure-property-function relationships of spider silks. Although in nature only fibers are formed from spidroins, in vitro, scientists can explore non-natural morphologies including nanofibrils, particles, capsules, hydrogels, films or foams. The versatility of spidroins, along with their biocompatible and biodegradable nature, also placed them as leading-edge biological macromolecules for improved drug delivery and various biomedical applications. Accordingly, in this review, we highlight the relationship between the molecular structure of spider silk and its mechanical properties and aims to provide a critical summary of recent progress in research employing recom...
Discussion Initially transgenic goats were utilized to produce spidersilk protein MaSp1 and MaSp2... more Discussion Initially transgenic goats were utilized to produce spidersilk protein MaSp1 and MaSp2. Our goal of large scale protein production and purification could not be met ideally by this method. It is very time consuming and inconvenient. For example in E. Coli, it is relatively easy to make multiple constructs but for a goat it would take a long time. Goats also cost more money to maintain (food, land, resources). Use of Spidersilk protein commercially would be more difficult using a transgenic animal such as goats due to regulations and negative stigma toward GMOs, as opposed to mass producing via E. Coli. Using bacteria such as E. Coli is beneficial due to its much lower cost margin, short generation time, and well understood and researched methods of inserting genes and producing desired proteins. Methods of Protein purification Our proteins MaSp1 and MaSp2 have a polyhistidine tag, which is a string of histidine residues typically at either N terminus or the C terminus of ...
Of the 7 silks produced; Major Ampullate silk is the strongest natural fiber known to man. Dragli... more Of the 7 silks produced; Major Ampullate silk is the strongest natural fiber known to man. Dragline spider silk is unique because of its strength, stretch and biocompatibility. These properties could allow it to be used to create diverse materials such as stronger cables, ultra fine sutures for eye and nerve surgeries, fire-proof clothing, etc. In order to manufacture these materials, one must be able to produce and purify the synthetic spider silk proteins that make up dragline silk, called Masp 1 (M4) and Masp 2 (M5). Dr. Lewis’s lab at USU has made multiple advancements in producing spider silk proteins. Some methods include fermenting E. coli, growing genetically modified silkworms, and raising alfalfa. One of the best ways the Lewis lab produces synthetic spider silk is in the mammary (milk) of dairy goats. The transgenic goats produce more protein than the other alternative synthetic production methods and it is the most effective and inexpensive purification method using a Ta...
Spider silk is one of the most robust and versa7le fibers making it a topic of interest in the sc... more Spider silk is one of the most robust and versa7le fibers making it a topic of interest in the scien7fic community. Possessing strength and elas7city many have sought to create fibers comparable to natural spider silk. Up un7l recently many scien7sts have fallen short of crea7ng said fibers. With the use of a technique called electrospinning, comparable spider silk fibers have been created. Electrospinning is the process of crea7ng fibers from a polymer solu7on using an electrical field. This method leads to the forma7on of nanofibers. These Fibers can then be further modified by crosslinking, a technique tradi7onally used to analyze protein-‐protein interac7ons. Introduc7on
Spider silk has impressive physical properties in the areas of strength, weight, toughness and el... more Spider silk has impressive physical properties in the areas of strength, weight, toughness and elasticity, with commercial use currently limited as large-scale production has not been demonstrated. Transgenic work to synthetically replicate the spider silk proteins via E. coli has successfully been performed. Spider silk based products show promising characteristics, but the economic viability of synthetic spider silk production has not been evaluated. This work presents an analysis of the economic viability and environmental impact associated with the production of synthetic spider silk through E. coli fermentation. System models, validated with experimental data, were leveraged with economic and life cycle modeling to evaluate the sustainability of synthetic spider silk production. Results show protein yield, modeling scale, and processing technologies dramatically impact final silk protein cost as well as the global warming potential.
Spider silk, an ancient biomaterial, has many qualities worth replicating. With the use of geneti... more Spider silk, an ancient biomaterial, has many qualities worth replicating. With the use of genetic modification, relatively large amounts of spider silk protein have been produced utilizing transgenic goat milk. Access to this protein has allowed us to create spider silk films and hydrogels. Applications for these products are far reaching, from football helmets to drug release capsules. Using mammalian cells, it has been proven that spider silk is not only biocompatible, but that it degrades at an ideal rate. As medical advances have taken place, the ability to create scaffolds for biomedical research and tissue implantation have increased. Spider silk films and hydrogels are an ideal material to consider for such scaffolds. This project focuses on discovering and improving mammalian cell growth on spider silk films and hydrogels so as to create optimal scaffolds. We plan to see beneficial cell growth and attachment by virtue of the composition of our spider silk films and hydrogels compared to our controls
Spider silk, which has remarkable mechanical properties, is a natural protein fiber produced by s... more Spider silk, which has remarkable mechanical properties, is a natural protein fiber produced by spiders. Spiders cannot be farmed because of their cannibalistic and territorial nature. Hence, large amounts of spider silk cannot be produced from spiders. Genetic engineering is an alternative approach to produce large quantities of spider silk. Our group has produced synthetic spider silk proteins in E.coli to study structure/function and to produce biomaterials comparable to the silks produced by orb-weaving spiders. Here we give a detailed description of our cloning, expression, and purification methods of synthetic spider silk proteins ranging from ∼30 to ∼200 kDa. We have cloned the relevant genes of the spider Nephila clavipes and introduced them into bacteria to produce synthetic spider silk proteins using small and large-scale bioreactors. We have optimized the fermentation process, and we have developed protein purification methods as well. The purified proteins are spun into fibers and are used to make alternative materials like films and adhesives with various possible commercial applications.
Spider silks are well known for their extraordinary mechanical properties. This characteristic is... more Spider silks are well known for their extraordinary mechanical properties. This characteristic is a result of the interplay of composition, structure and self-assembly of spider silk proteins (spidroins). Advances in synthetic biology have enabled the design and production of spidroins with the aim of biomimicking the structure-property-function relationships of spider silks. Although in nature only fibers are formed from spidroins, in vitro, scientists can explore non-natural morphologies including nanofibrils, particles, capsules, hydrogels, films or foams. The versatility of spidroins, along with their biocompatible and biodegradable nature, also placed them as leading-edge biological macromolecules for improved drug delivery and various biomedical applications. Accordingly, in this review, we highlight the relationship between the molecular structure of spider silk and its mechanical properties and aims to provide a critical summary of recent progress in research employing recom...
Discussion Initially transgenic goats were utilized to produce spidersilk protein MaSp1 and MaSp2... more Discussion Initially transgenic goats were utilized to produce spidersilk protein MaSp1 and MaSp2. Our goal of large scale protein production and purification could not be met ideally by this method. It is very time consuming and inconvenient. For example in E. Coli, it is relatively easy to make multiple constructs but for a goat it would take a long time. Goats also cost more money to maintain (food, land, resources). Use of Spidersilk protein commercially would be more difficult using a transgenic animal such as goats due to regulations and negative stigma toward GMOs, as opposed to mass producing via E. Coli. Using bacteria such as E. Coli is beneficial due to its much lower cost margin, short generation time, and well understood and researched methods of inserting genes and producing desired proteins. Methods of Protein purification Our proteins MaSp1 and MaSp2 have a polyhistidine tag, which is a string of histidine residues typically at either N terminus or the C terminus of ...
Of the 7 silks produced; Major Ampullate silk is the strongest natural fiber known to man. Dragli... more Of the 7 silks produced; Major Ampullate silk is the strongest natural fiber known to man. Dragline spider silk is unique because of its strength, stretch and biocompatibility. These properties could allow it to be used to create diverse materials such as stronger cables, ultra fine sutures for eye and nerve surgeries, fire-proof clothing, etc. In order to manufacture these materials, one must be able to produce and purify the synthetic spider silk proteins that make up dragline silk, called Masp 1 (M4) and Masp 2 (M5). Dr. Lewis’s lab at USU has made multiple advancements in producing spider silk proteins. Some methods include fermenting E. coli, growing genetically modified silkworms, and raising alfalfa. One of the best ways the Lewis lab produces synthetic spider silk is in the mammary (milk) of dairy goats. The transgenic goats produce more protein than the other alternative synthetic production methods and it is the most effective and inexpensive purification method using a Ta...
Spider silk is one of the most robust and versa7le fibers making it a topic of interest in the sc... more Spider silk is one of the most robust and versa7le fibers making it a topic of interest in the scien7fic community. Possessing strength and elas7city many have sought to create fibers comparable to natural spider silk. Up un7l recently many scien7sts have fallen short of crea7ng said fibers. With the use of a technique called electrospinning, comparable spider silk fibers have been created. Electrospinning is the process of crea7ng fibers from a polymer solu7on using an electrical field. This method leads to the forma7on of nanofibers. These Fibers can then be further modified by crosslinking, a technique tradi7onally used to analyze protein-‐protein interac7ons. Introduc7on
Spider silk has impressive physical properties in the areas of strength, weight, toughness and el... more Spider silk has impressive physical properties in the areas of strength, weight, toughness and elasticity, with commercial use currently limited as large-scale production has not been demonstrated. Transgenic work to synthetically replicate the spider silk proteins via E. coli has successfully been performed. Spider silk based products show promising characteristics, but the economic viability of synthetic spider silk production has not been evaluated. This work presents an analysis of the economic viability and environmental impact associated with the production of synthetic spider silk through E. coli fermentation. System models, validated with experimental data, were leveraged with economic and life cycle modeling to evaluate the sustainability of synthetic spider silk production. Results show protein yield, modeling scale, and processing technologies dramatically impact final silk protein cost as well as the global warming potential.
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