Triggered Morphology Generation in a Biosynthetic Model Spider Dragline Silk Protein

Recombinant DNA techniques were used to prepare a protein modeled after the consensus sequence of Nephila clavipesspider dragline silk, incorporating methionine residues to serve as redox “triggers”. In addition a water-soluble 27 residue peptide model of the dragline silk consensus amorphous sequence, representing a single amorphous block in the protein sequence, was prepared and characterized to gain additional insight into the behavior of the amorphous phase. X-ray diffraction, electron diffraction, transmission electron microscopy (TEM), and Fourier transform infrared spectroscopy (FTIR) were used to characterize the ability of the recombinant protein to form (β-sheet crystals and the effect of the oxidation state of the redox trigger on crystallinity and noncrystalline order in the sample. The formation of intractable β-sheet crystallites is a major cause of insolubility in proteins that can form this type of secondary structure. Changes in crystallinity were observed when triggered/reduced (insoluble) and untriggered/oxidized (soluble) protein samples were compared.

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Packing Structure of Antiparallel β-Sheet Polyalanine Region in a Sequential Model Peptide of Nephila clavipes Dragline Silk Studied Using 13C Solid-State NMR and MD Simulation

Biomacromolecules ◽

10.1021/acs.biomac.9b00969 ◽

2019 ◽

Vol 20 (10) ◽

pp. 3884-3894 ◽

Cited By ~ 1

Author(s):

Tetsuo Asakura ◽

Akio Nishimura ◽

Akihiro Aoki ◽

Akira Naito

Keyword(s):

Solid State ◽

Solid State Nmr ◽

Md Simulation ◽

Nephila Clavipes ◽

Model Peptide ◽

Dragline Silk ◽

Sequential Model ◽

Packing Structure ◽

Β Sheet

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Structure of the N-terminal domain of Euprosthenops australis dragline silk suggests that conversion of spidroin dope to spider silk involves a conserved asymmetric dimer intermediate

Acta Crystallographica Section D Structural Biology ◽

10.1107/s2059798319007253 ◽

2019 ◽

Vol 75 (7) ◽

pp. 618-627 ◽

Cited By ~ 1

Author(s):

Wangshu Jiang ◽

Glareh Askarieh ◽

Alexander Shkumatov ◽

My Hedhammar ◽

Stefan D. Knight

Keyword(s):

Electrostatic Interactions ◽

Spider Silk ◽

Quaternary Structure ◽

Nephila Clavipes ◽

Trigger Mechanism ◽

Dragline Silk ◽

Terminal Domain ◽

Structure Changes ◽

Molecular Events ◽

Asymmetric Dimer

Spider silk is a biomaterial with exceptional mechanical toughness, and there is great interest in developing biomimetic methods to produce engineered spider silk-based materials. However, the mechanisms that regulate the conversion of spider silk proteins (spidroins) from highly soluble dope into silk are not completely understood. The N-terminal domain (NT) of Euprosthenops australis dragline silk protein undergoes conformational and quaternary-structure changes from a monomer at a pH above 7 to a homodimer at lower pH values. Conversion from the monomer to the dimer requires the protonation of three conserved glutamic acid residues, resulting in a low-pH `locked' dimer stabilized by symmetric electrostatic interactions at the poles of the dimer. The detailed molecular events during this transition are still unresolved. Here, a 2.1 Å resolution crystal structure of an NT T61A mutant in an alternative, asymmetric, dimer form in which the electrostatic interactions at one of the poles are dramatically different from those in symmetrical dimers is presented. A similar asymmetric dimer structure from dragline silk of Nephila clavipes has previously been described. It is suggested that asymmetric dimers represent a conserved intermediate state in spider silk formation, and a revised `lock-and-trigger' mechanism for spider silk formation is presented.

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Nanostructure and molecular mechanics of spider dragline silk protein assemblies

Journal of The Royal Society Interface ◽

10.1098/rsif.2010.0149 ◽

2010 ◽

Vol 7 (53) ◽

pp. 1709-1721 ◽

Cited By ~ 156

Author(s):

Sinan Keten ◽

Markus J. Buehler

Keyword(s):

Secondary Structure ◽

Spider Silk ◽

Mechanical Performance ◽

Experimental Studies ◽

Silk Protein ◽

Nephila Clavipes ◽

Protein Assemblies ◽

Dragline Silk ◽

Spider Dragline Silk ◽

Secondary Structure Content

Spider silk is a self-assembling biopolymer that outperforms most known materials in terms of its mechanical performance, despite its underlying weak chemical bonding based on H-bonds. While experimental studies have shown that the molecular structure of silk proteins has a direct influence on the stiffness, toughness and failure strength of silk, no molecular-level analysis of the nanostructure and associated mechanical properties of silk assemblies have been reported. Here, we report atomic-level structures of MaSp1 and MaSp2 proteins from the Nephila clavipes spider dragline silk sequence, obtained using replica exchange molecular dynamics, and subject these structures to mechanical loading for a detailed nanomechanical analysis. The structural analysis reveals that poly-alanine regions in silk predominantly form distinct and orderly beta-sheet crystal domains, while disorderly regions are formed by glycine-rich repeats that consist of 3 1 -helix type structures and beta-turns. Our structural predictions are validated against experimental data based on dihedral angle pair calculations presented in Ramachandran plots, alpha-carbon atomic distances, as well as secondary structure content. Mechanical shearing simulations on selected structures illustrate that the nanoscale behaviour of silk protein assemblies is controlled by the distinctly different secondary structure content and hydrogen bonding in the crystalline and semi-amorphous regions. Both structural and mechanical characterization results show excellent agreement with available experimental evidence. Our findings set the stage for extensive atomistic investigations of silk, which may contribute towards an improved understanding of the source of the strength and toughness of this biological superfibre.

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Nephila Clavipes Dragline Silk: Approaches to a Recombinantly Produced Silk Protein

MRS Proceedings ◽

10.1557/proc-330-37 ◽

1993 ◽

Vol 330 ◽

Author(s):

Charlene M. Mello ◽

Steven Arcidiacono ◽

Richard Beckwitt ◽

John Prince ◽

Kris Senecal ◽

...

Keyword(s):

High Performance ◽

Spider Silk ◽

Recombinant Dna ◽

Electromagnetic Spectrum ◽

Nephila Clavipes ◽

Natural Function ◽

High Performance Fiber ◽

Recombinant Dna Techniques ◽

Clone And Express ◽

Spider Silks

Spider silks exhibit an unusual combination of strength and toughness that distinguishes them from other natural and synthetic fibers. Silk proteins perform a key natural function as structural fibers, to absorb impact energy from flying insects without breaking. They dissipate energy over a broad area and balance stiffness, strength and extensibility (1,2). In addition to their unusual mechanical properties and visual lustre, silks also exhibit interesting interference patterns within the electromagnetic spectrum (3), unusual viscometric patterns related to processing (4), and piezoelectric properties (3,5,6). These properties suggest they would be good candidates for high performance fiber and composite applications. However, the spider is not capable of producing sufficient quantities of proteins to enable thorough evaluation of their potential. Consequently, we are pursuing recombinant DNA techniques to clone and express adequate quantities of recombinant spider silk for these studies.

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Scalable Spider Silk Inspired Materials with High Extensibility and Super Toughness

Key Engineering Materials ◽

10.4028/www.scientific.net/kem.893.31 ◽

2021 ◽

Vol 893 ◽

pp. 31-35

Author(s):

Jin Lian Hu ◽

Yuan Zhang Jiang ◽

Lin Gu

Keyword(s):

Chemical Synthesis ◽

Spider Silk ◽

Helical Structure ◽

Silk Proteins ◽

Dragline Silk ◽

Spider Dragline Silk ◽

Argiope Trifasciata ◽

Chemical Synthesis Route ◽

Β Sheet ◽

Huge Challenge

Spiders silks have extraordinary strength and toughness simultaneously, thus has become dreamed materials by scientists and industries. Although there have been tremendous attempts to prepare fibers from genetically manufacture spider silk proteins, however, it has been still a huge challenge because of tedious procedure and high cost. Here, a facile spider-silk-mimicking strategy is reported for preparing highly scratchable polymers and supertough fibers from chemical synthesis route. Polymer films with high extensibility (>1200%) and supertough fibers (~387 MJ m-3) are achieved by introducing polypeptides with β-sheet and α-helical structure in polyureathane/urea polymers. Notabley,the toughness of the fiber is more than twice the reported value of a normal spider dragline silk, and comparable with the toughest spider silk, aciniform silk of Argiope trifasciata.

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