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Identification of an aqueous glue protein, SCP-2, and the development of a polyclonal antiserum against the bHLH transcription factor SGSF in Latrodectus Hesperus
Date of Award
Thesis - Pacific Access Restricted
Master of Science (M.S.)
Craig A. Vierra
First Committee Member
Lisa A. Wrischnik
Second Committee Member
Although numerous spider fibroins have been reported, no known silk coating peptides have been discovered. We provide the first biochemical evidence for a spider coating peptide, called SCP-2, found on gumfooted lines, scaffolding joints and egg cases. The presence of this spider coating peptide on the fibers is supported by MS/MS analysis. Using quantitative real-time PCR analysis, we also demonstrate that SCP-2 has a flagelliform-restricted mRNA pattern of expression. Molecular modeling of the SCP-2 amino acid sequence predicts it adopts an alpha-helical structure that is amphipathic in nature. SCP-2, which can be extracted from fibers using water, is hypothesized to influence the mechanical properties of the silk fibers as well as serve a protective function for the threads. Based upon the restricted pattern of expression of SCP-2, our findings reveal novel insight regarding the glandular function of the flagelliform gland in . cob weaving spiders, suggesting it produces aqueous coating materials that are deposited on a wide range of different silk types. In addition, in an attempt to advance our understanding regarding silk gene transcription, our lab has developed the first antibody against the bHLH factor SGSF. SGSF has been implicated as a potential transcriptional regulator of silk gene transcription in spiders. Development of the anti-SGSF antibody was accomplished via the overexpression and purification of a fusion protein in bacteria, which consisted of the C-terminal region of SGSF fused to thioredoxin. Purified SGSF fusion proteins were injected into rabbits and the polyclonal antiserum was collected and tested by western blot analysis to determine the specificity of the immunological reagent. Western blot analyses revealed the anti-SGSF antiserum was capable of recognizing bacterially expressed SGSF in an efficient manner. Collectively, these studies lay the groundwork for future investigations involving the use of the antibody to determine the role of SGSF in silk transcription.
La Mattina, Coby Ann. (2009). Identification of an aqueous glue protein, SCP-2, and the development of a polyclonal antiserum against the bHLH transcription factor SGSF in Latrodectus Hesperus. University of the Pacific, Thesis - Pacific Access Restricted. https://scholarlycommons.pacific.edu/uop_etds/716
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