INTELLECTUAL MERIT: The goal of this project is to investigate structure-function relationships in the spider protein building blocks that are responsible for the transformation of soluble silk into insoluble fibers with unrivaled mechanical properties. Spider dragline silk consists almost exclusively of two large proteins (major ampullate spidroins, MaSp1 and MaSp2) that contain short, non-repetitive N- and C-terminal domains (NTD and CTD, respectively) flanking a large, highly repetitive central domain. Dragline silk has unmatched toughness combined with elasticity. It presents a demanding target for attempts at biomimetic reproduction of these properties. In order to exploit the potential of natural protein-based fibers for new biomaterials, it will be necessary to understand fully the mechanism and underlying principles of dragline fiber self-assembly. To test the hypothesis that pH- and salt-dependent differences in the structures of the N-terminal domain contribute to the transition of soluble protein in the lumen of the spinning gland into insoluble dragline fiber, the PIs will (1) determine the structures of MaSp-NTD under conditions mimicking spider gland and duct environments. They propose that a pH-induced conformational change has evolved as a mechanism for sensing the drop in pH that occurs along the length of the major ampullate duct and functions as an important signal in the fiber self-assembly process. Thus, the PIs will (2) analyze the MaSp-NTD dimer interface and determine the mechanism of pH sensing. Finally, to test the hypothesis that the role of MaSp-NTD dimerization is to promote the formation of long multimeric strands of spidroin molecules and that these long multimeric strands then assemble upon one another to form a fiber, these studies aim (3) to clarify the functional role of MaSp-NTD in the context of mini-silk constructs.

BROADER IMPACTS: These protein structure-function studies have potentially far-reaching implications. Understanding protein conformational transitions associated with highly soluble, monomeric states and insoluble, oligomeric states promises to shed light on general properties of proteins. This information will serve not only to guide the development of new protein-based biomimetic materials but may also provide insights into the assembly process that can be used with non-proteinaceous polymers. In addition, as we learn more about the basic biology of protein assembly, the principles may be applied to other proteins that aggregate to form fibrils. Some of these are associated with protein misfolding and its dangerous consequences in the context of amyloidosis. This project opens new opportunities for fundamental studies in the relationship between protein structure and function and spider silk biomimetic manufacturing. The proposed research reaches across disciplines and two institutions of higher education in South Carolina. The Medical University of South Carolina (MUSC) and Clemson University (CU) have a long history of cooperation, and this inter-institutional project strengthens the collaborative research and education programs of both universities. The diverse and interdisciplinary research focusing on properties of spider silk proteins provides a rich learning environment for postdoctoral fellows, graduate students, and fertile training grounds for undergraduates participating in research activities at CU and in MUSC's Summer Undergraduate Research Program (SURP). Students will be exposed to ongoing, cutting-edge NMR research that is integrated within the Center for Structural Biology at MUSC. The project provides support for our key outreach effort, the bi-annual Charleston NMR Summer Camp at the Hollings Marine Laboratory (HML). The Camp's long-term goal is to attract and develop the next generation of scientists with a passion for cutting-edge NMR in South Carolina, with special emphasis on the state's four-year colleges and minority-serving institutions.

National Science Foundation (NSF)
Division of Materials Research (DMR)
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mohan srinivasarao
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Medical University of South Carolina
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