Title

Biochemical Expression and Characterization of CRISP3, a Novel Protein Found in Spider Dragline Silk Fibers

Poster Number

01B

Lead Author Major

Biochemistry with Empahsis in Pre-Medicine

Lead Author Status

Senior

Format

Poster Presentation

Faculty Mentor Name

Craig Vierra

Faculty Mentor Email

cvierra@pacific.edu

Faculty Mentor Department

Biological Department

Abstract/Artist Statement

Spider silk represents an outstanding biomaterial spun from protein. It can outperform a large number of manmade materials, including high tensile steel and Kevlar™ (body armor). Moreover, spider silk also displays antimicrobial activity and is a green material. Spiders can spin 6-7 different fiber types with distinct biological properties. One of the fiber types, dragline silk, is used for locomotion and web construction. This fiber type contains at least seven distinct proteins: three members of the Cysteine-Rich Protein Family (CRP1, CRP2 and CRP4), CRISP3, fasciclin and two uncharacterized proteins. Our research study focuses on the molecular characterization of CRISP3. In these studies, we amplified the cDNA coding for CRISP3 from a library prepared from silk-producing glands of the black widow spider. The amplified product was inserted into a prokaryotic expression vector. After heterologous expression of CRISP3 in bacteria, we purified the recombinant protein using affinity chromatography. Purification of CRISP3 was confirmed by performing liquid chromatography mass spectrometry (LC-MS) analysis using an Orbitrap Fusion™ Tribrid™ mass spectrometer. Our long-term goal is to use purified CRISP3 and mass spectrometry to study its three-dimensional structure. These data are important to help elucidate the assembly processes of dragline silk, ultimately facilitating the development of new synthetic spider silk materials for industrial applications.

Location

DeRosa University Center, Ballroom

Start Date

29-4-2017 1:00 PM

End Date

29-4-2017 3:00 PM

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Apr 29th, 1:00 PM Apr 29th, 3:00 PM

Biochemical Expression and Characterization of CRISP3, a Novel Protein Found in Spider Dragline Silk Fibers

DeRosa University Center, Ballroom

Spider silk represents an outstanding biomaterial spun from protein. It can outperform a large number of manmade materials, including high tensile steel and Kevlar™ (body armor). Moreover, spider silk also displays antimicrobial activity and is a green material. Spiders can spin 6-7 different fiber types with distinct biological properties. One of the fiber types, dragline silk, is used for locomotion and web construction. This fiber type contains at least seven distinct proteins: three members of the Cysteine-Rich Protein Family (CRP1, CRP2 and CRP4), CRISP3, fasciclin and two uncharacterized proteins. Our research study focuses on the molecular characterization of CRISP3. In these studies, we amplified the cDNA coding for CRISP3 from a library prepared from silk-producing glands of the black widow spider. The amplified product was inserted into a prokaryotic expression vector. After heterologous expression of CRISP3 in bacteria, we purified the recombinant protein using affinity chromatography. Purification of CRISP3 was confirmed by performing liquid chromatography mass spectrometry (LC-MS) analysis using an Orbitrap Fusion™ Tribrid™ mass spectrometer. Our long-term goal is to use purified CRISP3 and mass spectrometry to study its three-dimensional structure. These data are important to help elucidate the assembly processes of dragline silk, ultimately facilitating the development of new synthetic spider silk materials for industrial applications.