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Molecular mechanisms in silk production and biomimetic spider silk spinning

Anna Rising

Jeudi 11 mai

à 11h, en salle C. BROT

In order to mimic spider silk formation, the complex processes that govern the assembly of spider silk proteins (spidroins) must be understood. We have studied the spider silk glands extensively and found a highly regulated pH gradient along the glands. Molecular studies of recombinant spidroins under the pHs found in the glands revealed a lock and trigger mechanism that control spider silk formation [1]. The N-terminal domain (NT) is the most conserved part of spider silk proteins (spidroins) and dimerizes and gets stabilized in a pH dependent three-step mechanism, while the C-terminal domain (CT) undergoes unfolding and assembles into amyloid-like fibrils [2, 3]. These molecular events secure high spidroin solubility under storage conditions, promote stable NT dimerization that locks spidroins into an end-less network during silk formation, and allow CT to trigger -sheet formation that make up the stable silk fibres. We realized that terminal domains from different silks and species have different solubility when expressed in E.coli. Based on these insights we have designed novel recombinant spidroins that are expressed at large yields, are soluble at >500mg/mL at native conformations and that respond to changes in pH according to the lock and trigger mechanism. To mimic the pH gradient of the gland we built a spinning device from pulled glass capillaries that enables water based spinning of recombinant spidroins into “end-less” fibers, and we can now, for the first time, truly recapitulate the spinning of spider silk in vitro [4].

References :

1. Rising, A. Johansson, J. (2015) Toward spinning artificial spider silk. Nat Chem Biol. 11, (5), 309-15.

2. Andersson, M., Chen, G., Otikovs, M., Landreh, M., Nordling, K., Kronqvist, N., Westermark, P., Jörnvall, H., Knight, S.D., Ridderstråle, Y., Holm, L., Meng, Q., Jaudzems, K., Chesler, M., Johansson, J. Rising, A. (2014) Carbonic anhydrase generates CO2 and H+ that drive spider silk formation via opposite effects on the terminal domains. PLoS Biol. 12, e1001921.

3. Kronqvist, N., Otikovs, M., Chmyrov, V., Chen, G., Andersson, M., Nordling, K., Landreh, M., Sarr, M., Jornvall, H., Wennmalm, S., Widengren, J., Meng, Q., Rising, A., Otzen, D., Knight, S.D., Jaudzems, K. Johansson, J. (2014) Sequential pH-driven dimerization and stabilization of the N-terminal domain enables rapid spider silk formation. Nat Commun. 5, 3254.

4. Andersson, M., Jia, Q., Abella, A., Lee, X.Y., Landreh, M., Purhonen, P., Hebert, H., Tenje, M., Robinson, C.V., Meng, Q., Plaza, G.R., Johansson, J. Rising, A. (2017) Biomimetic spinning of artificial spider silk from a chimeric minispidroin. Nat Chem Biol. 13, (3), 262-264.