Spider Silk Fibers Spun from Soluble Recombinant Silk Produced in Mammalian Cells

Spider silks are protein-based “biopolymer” filaments or threads secreted by specialized epithelial cells as concentrated soluble precursors of highly repetitive primary sequences. Spider dragline silk is a flexible, lightweight fiber of extraordinary strength and toughness comparable to that of synthetic high-performance fibers. We sought to “biomimic” the process of spider silk production by expressing in mammalian cells the dragline silk genes (ADF-3/MaSpII and MaSpI) of two spider species. We produced soluble recombinant (rc)–dragline silk proteins with molecular masses of 60 to 140 kilodaltons. We demonstrated the wet spinning of silk monofilaments spun from a concentrated aqueous solution of soluble rc–spider silk protein (ADF-3; 60 kilodaltons) under modest shear and coagulation conditions. The spun fibers were water insoluble with a fine diameter (10 to 40 micrometers) and exhibited toughness and modulus values comparable to those of native dragline silks but with lower tenacity. Dope solutions with rc–silk protein concentrations >20% and postspinning draw were necessary to achieve improved mechanical properties of the spun fibers. Fiber properties correlated with finer fiber diameter and increased birefringence.

[1]  L. Bedzyk,et al.  Microbial production of spider silk proteins. , 2000, Journal of biotechnology.

[2]  R. Lewis,et al.  Extreme Diversity, Conservation, and Convergence of Spider Silk Fibroin Sequences , 2001, Science.

[3]  D. Ginzinger,et al.  Silk Properties Determined by Gland-Specific Expression of a Spider Fibroin Gene Family , 1996, Science.

[4]  Robert W. Work,et al.  Dimensions, Birefringences, and Force-Elongation Behavior of Major and Minor Ampullate Silk Fibers from Orb-Web-Spinning Spiders—The Effects of Wetting on these Properties , 1977 .

[5]  Mark W. Denny,et al.  THE PHYSICAL PROPERTIES OF SPIDER'S SILK AND THEIR ROLE IN THE DESIGN OF ORB-WEBS , 1976 .

[6]  Manuel Elices,et al.  Structural biological materials : design and structure-property relationships , 2000 .

[7]  Oskar Liivak,et al.  Artificial Spinning of Spider Silk , 1998 .

[8]  R. Lewis,et al.  Hypotheses that correlate the sequence, structure, and mechanical properties of spider silk proteins. , 1999, International journal of biological macromolecules.

[9]  G C Candelas,et al.  Spider silkglands contain a tissue-specific alanine tRNA that accumulates in vitro in response to the stimulus for silk protein synthesis. , 1990, Developmental biology.

[10]  R. Lewis,et al.  Molecular architecture and evolution of a modular spider silk protein gene. , 2000, Science.

[11]  Fritz Vollrath,et al.  Liquid crystalline spinning of spider silk , 2001, Nature.

[12]  D. Kaplan,et al.  Construction, cloning, and expression of synthetic genes encoding spider dragline silk. , 1995, Biochemistry.

[13]  P. Yager,et al.  Comparative Structural Characterization of Naturally- and Synthetically-Spun Fibers of Bombyx mori Fibroin , 1998 .

[14]  J. Scheller,et al.  Production of spider silk proteins in tobacco and potato , 2001, Nature Biotechnology.

[15]  Oskar Liivak,et al.  A Microfabricated Wet-Spinning Apparatus To Spin Fibers of Silk Proteins. StructureProperty Correlations , 1998 .

[16]  P. Lizardi,et al.  Discontinuous translation of silk fibroin in a reticulocyte cell-free system and in intact silk gland cells. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[17]  L. Jelinski,et al.  Regenerated Spider Silk: Processing, Properties, and Structure , 2000 .

[18]  M B Hinman,et al.  Isolation of a clone encoding a second dragline silk fibroin. Nephila clavipes dragline silk is a two-protein fiber. , 1992, The Journal of biological chemistry.

[19]  R. Lewis,et al.  Expression and purification of a spider silk protein: a new strategy for producing repetitive proteins. , 1996, Protein expression and purification.

[20]  S. Fahnestock,et al.  Synthetic spider dragline silk proteins and their production in Escherichia coli , 1997, Applied Microbiology and Biotechnology.

[21]  R. Lewis,et al.  Structure of a protein superfiber: spider dragline silk. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[22]  Robert W. Work,et al.  The Force-Elongation Behavior of Web Fibers and Silks Forcibly Obtained from Orb-Web-Spinning Spiders , 1976 .

[23]  J. Gosline,et al.  The mechanical design of spider silks: from fibroin sequence to mechanical function. , 1999, The Journal of experimental biology.

[24]  M B Hinman,et al.  Synthetic spider silk: a modular fiber. , 2000, Trends in biotechnology.

[25]  D. Kaplan,et al.  Purification and characterization of recombinant spider silk expressed in Escherichia coli , 1998, Applied Microbiology and Biotechnology.

[26]  L. Bedzyk,et al.  Production of synthetic spider dragline silk protein in Pichia pastoris , 1997, Applied Microbiology and Biotechnology.

[27]  W. Nentwig Ecophysiology of Spiders , 1987, Springer Berlin Heidelberg.