Degree of Biomimicry of Artificial Spider Silk Spinning Assessed by NMR Spectroscopy

Otikovs, Mārtiņš; Andersson, Marlene; Jia, Qiupin; Nordling, Kerstin; Meng, Qing; Andreas, Loren B.; Pintacuda, Guido; Johansson, Jan; Rising, Anna; Jaudzems, Kristaps

doi:10.1002/ange.201706649

Cited by 11 publications

(11 citation statements)

References 42 publications

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“… 44 NT2RepCT in solution also forms assemblies of 4–6 molecules, which are around 7–11 nm in diameter. 29 , 30 In this study, we observed that the shear viscosity at 1 s –1 of NT2RepCT ( Figure 1 A), or the consistency index from fitting the Herschel–Bulkley model ( Figure 2 C) correlated with the concentration. This suggests that the NT2RepCT micelles interact, and in line with this, increased concentration of NT2RepCT leads to increased viscosity of the dope.…”

Section: Discussionsupporting

confidence: 50%

“… 8 , 46 , 48 , 53 − 56 By using CD spectroscopy, we found that NT2RepCT is mainly in an α-helical conformation at pH 8.0, 7.0, and 6.5 ( Figure 4 A, B and Figure S3B ), which is expected from the α-helical terminal domains and predicted α-helical conformation of the poly alanine regions, and also in line with previous findings where NT2RepCT was studied by NMR spectroscopy. 30 The extreme solubility of NT2RepCT enabled us to study this protein in solution by FTIR spectroscopy, which confirmed that, even at high concentrations, the protein conformation was dominated by α-helical and β-turn components ( Figure 5 ). Previous FTIR spectroscopy studies on gland content also suggested the presence of α-helices accompanied by unidentified secondary structures.…”

Section: Discussionmentioning

confidence: 59%

“…The assembly of NT2RepCT is pH dependent and recapitulates important molecular mechanisms associated with native silk spinning. 29 , 30 …”

Section: Introductionmentioning

confidence: 99%

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Native-like Flow Properties of an Artificial Spider Silk Dope

Arndt

Laity

Johansson

et al. 2021

ACS Biomater. Sci. Eng.

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Recombinant spider silk has emerged as a biomaterial that can circumvent problems associated with synthetic and naturally derived polymers, while still fulfilling the potential of the native material. The artificial spider silk protein NT2RepCT can be produced and spun into fibers without the use of harsh chemicals and here we evaluate key properties of NT2RepCT dope at native-like concentrations. We show that NT2RepCT recapitulates not only the overall secondary structure content of a native silk dope but also emulates its viscoelastic rheological properties. We propose that these properties are key to biomimetic spinning and that optimization of rheological properties could facilitate successful spinning of artificial dopes into fibers.

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Section: Discussionsupporting

confidence: 50%

Section: Discussionmentioning

confidence: 59%

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Native-like Flow Properties of an Artificial Spider Silk Dope

Arndt

Laity

Johansson

et al. 2021

ACS Biomater. Sci. Eng.

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“…4E). This is particularly 40 shows that the fusion protein is located in inclusion bodies. A representative example from two repeats is shown.…”

Section: Fibril Formation Mediated By An Exposed Amyloidogenic Sequence Does Not Results In Global Nt Aggregationmentioning

confidence: 90%

High intracellular stability of the spidroin N‐terminal domain in spite of abundant amyloidogenic segments revealed by in‐cell hydrogen/deuterium exchange mass spectrometry

et al. 2019

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Proteins require an optimal balance of conformational flexibility and stability in their native environment to ensure their biological functions. A striking example is spidroins, spider silk proteins, which are stored at extremely high concentrations in soluble form, yet undergo amyloid‐like aggregation during spinning. Here, we elucidate the stability of the highly soluble N‐terminal domain (NT) of major ampullate spidroin 1 in the Escherichia coli cytosol as well as in inclusion bodies containing fibrillar aggregates. Surprisingly, we find that NT, despite being largely composed of amyloidogenic sequences, showed no signs of concentration‐dependent aggregation. Using a novel intracellular hydrogen/deuterium exchange mass spectrometry (HDX‐MS) approach, we reveal that NT adopts a tight fold in the E. coli cytosol and in this manner conceals its aggregation‐prone regions by maintaining a tight fold under crowded conditions. Fusion of NT to the unstructured amyloid‐forming Aβ 40 peptide, on the other hand, results in the formation of fibrillar aggregates. However, HDX‐MS indicates that the NT domain is only partially incorporated into these aggregates in vivo . We conclude that NT is able to control its aggregation to remain functional under the extreme conditions in the spider silk gland.

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“… 12 Expression and purification of spidroins under benign conditions, as well as subsequent biomimetic spinning protocols, have been developed and shown to recapitulate important events of the assembly process. 13 − 16 However, the mechanical properties of fibers produced in this manner are still inferior to those of the native silk fiber, likely due to the small size of the spidroins used; hence, there is a need for novel ideas on how to produce high-performance spider silk replicas using biotechnological tools.…”

Section: Introductionmentioning

confidence: 99%

Doing What Spiders Cannot—A Road Map to Supreme Artificial Silk Fibers

Johansson

Rising

2021

ACS Nano

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Fabricating artificial spider silk fibers in bulk scale has been a major goal in materials science for centuries. Two main routes have emerged for making such fibers. One method uses biomimetics in which the spider silk proteins (spidroins) are produced under nativelike conditions and then spun into fibers in a process that captures the natural, complex molecular mechanisms. However, these fibers do not yet match the mechanical properties of native silk fibers, potentially due to the small size of the designed spidroin used. The second route builds on biotechnological progress that enables production of large spidroins that can be spun into fibers by using organic solvents. With this approach, fibers that equal the native material in terms of mechanical properties can be manufactured, but the yields are too low for economically sustainable production. Hence, the need for new ideas is urgent. Herein, we introduce a structural-biology-based approach for engineering artificial spidroins that circumvents the laws with which spidroins, being secretory proteins, have to comply in order to avoid membrane insertion and provide a road map to the production of biomimetic silk fibers with improved mechanical properties.

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Degree of Biomimicry of Artificial Spider Silk Spinning Assessed by NMR Spectroscopy

Cited by 11 publications

References 42 publications

Native-like Flow Properties of an Artificial Spider Silk Dope

Native-like Flow Properties of an Artificial Spider Silk Dope

High intracellular stability of the spidroin N‐terminal domain in spite of abundant amyloidogenic segments revealed by in‐cell hydrogen/deuterium exchange mass spectrometry

Doing What Spiders Cannot—A Road Map to Supreme Artificial Silk Fibers

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