IP Library › Granted Patent US 12,209,331
Granted Patent B2
US 12,209,331 · App. 17/527,617 · Granted Jan 28, 2025

Methods of generating highly-crystalline recombinant spider silk protein fibers

Inventors: Amir Ahmad Bakhtiary Davijani (Oakland, CA); Steven Tom (Berkeley, CA); Maxime Boulet-Audet (El Cerrito, CA); Paul Andre Guerette (Oakland, CA); Lindsay Wray (Benicia, CA); Nicole Elizabeth Subler (Berkeley, CA); Zoya Nasir Zulfiqar (Oakland, CA); Hua Li (Davis, CA); Jessica Wang (Oakland, CA)
Assignee: BOLT THREADS, INC.
D01D5/06D01D1/02D01F4/02D01F6/68D02J1/221D02J13/005
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Quick Facts
Patent No.
US 12,209,331
App. No.
17/527,617
Granted
Jan 28, 2025
Kind
B2
Abstract

Provided herein are scalable methods of processing wet-spun fiber comprising recombinant spider silk polypeptides to generate a three-dimensional crystalline lattice of beta-sheet structures in the fiber.

Claims (35)

1. A method for generating a drawn fiber comprising a silk polypeptide, the method comprising:

dissolving a powder comprising the silk polypeptide into a solvent to generate a spin dope;

extruding the spin dope into a coagulation bath to form a precursor fiber;

collecting the precursor fiber without drawing the precursor fiber;

annealing the precursor fiber prior to drawing the fiber; and

drawing the precursor fiber only over a surface having a temperature of at least 190° C. to generate a drawn fiber,

wherein the drawn fiber has a crystallinity index of at least 6% as measured using X-ray diffraction.

2. The method of claim 1 , wherein said silk polypeptide is a recombinant silk polypeptide.

3. The method of claim 2 , wherein said recombinant silk polypeptide comprises two or more repeat units of a proteinaceous block copolymer.

4. The method of claim 1 , wherein said silk polypeptide is a recombinant spider silk polypeptide.

5. The method of claim 4 , wherein the recombinant spider silk polypeptide comprises SEQ ID NO: 1.

6. The method of claim 1 , wherein the powder comprising the silk polypeptide is comprised of at least 60% silk polypeptide by weight.

7. The method of claim 1 , wherein the drawn fiber is drawn over the surface at a draw ratio of at least 2×.

8. The method of claim 7 , wherein the draw ratio is computed by determining the draw ratio at failure.

9. The method of claim 8 , wherein determining the draw ratio at failure comprises determining the distribution of maximum elongation at break of one or more precursor fibers using an apparatus designed to draw the fiber over a surface having a temperature of at least 190° C. while increasing the draw ratio.

10. The method of claim 1 , further comprising drawing the drawn fiber over the surface having a temperature of at least 190° C. one or more times.

11. The method of claim 10 , wherein the sum of the draw ratios at each drawing step is approximately equal to or less than the draw ratio at failure of the precursor fiber.

12. The method of claim 10 , wherein the generation of the drawn fiber comprises:

determining a draw ratio at failure of the precursor fiber; and

distributing the draw ratio at failure of the precursor fiber over each of said drawing steps.

13. The method of claim 1 , wherein annealing the precursor fiber comprises annealing the precursor fiber with alcohol vapor.

14. The method of claim 1 , wherein the solvent comprises N-methyl morpholine N-oxide (NMMO) or formic acid.

15. The method of claim 14 , wherein the solvent comprises 20% to 60% by weight NMMO.

16. The method of claim 14 , wherein generating said drawn fiber comprises heating said spin dope before said step of extruding the spin dope into said coagulation bath.

17. The method of claim 14 , wherein extruding the spin dope into said coagulation bath comprises extruding the spin dope through an air in the range of 2 to 20 cm.

18. The method of claim 1 , wherein the drawn fiber has increased beta-sheet formation relative to the precursor fiber.

19. The method of claim 18 , wherein the drawn fiber has increased beta-sheet formation relative to the precursor fiber proportional to the draw ratio used to draw the fiber over the surface.

20. The method of claim 1 , wherein the surface is at least 200 degrees Celsius.

21. The method of claim 1 , wherein the surface is at least 20 degrees Celsius greater than the glass transition temperature of the precursor fiber.

22. The method of claim 1 , wherein the tenacity of the drawn fiber is greater than 20 cN/tex.

23. The method of claim 1 , wherein the Herman orientation factor of the drawn fiber is approximately the same as native silk fiber.

24. The method of claim 1 , wherein the drawn fiber has a crystallinity index of at least 7% as measured using X-ray diffraction.

25. The method of claim 1 , wherein the drawn fiber has more than 1.5 times increased beta-sheet content as compared to air drawn fibers subject to identical drawing conditions except not drawn over a surface having a temperature of at least 190° C.

26. The method of claim 1 , wherein the drawn fiber has an increased 3D β-sheet content as compared to air drawn fibers subject to identical drawing conditions except not drawn over a surface having a temperature of at least 190° C.

27. The method of claim 1 , further comprising drying the precursor fiber after the collecting and before the annealing to form a dried precursor fiber, wherein the drawing comprises drawing the dried precursor fiber only over the surface to generate the drawn fiber.

Assignments (2)
SECURITY INTEREST Recorded Oct 14, 2022
From: BOLT THREADS, INC.
To: GINKGO BIOWORKS, INC.
Reel/Frame 061430/0311 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Nov 18, 2021
From: BAKHTIARY DAVIJANI, AMIR AHMAD; TOM, STEVEN; BOULET-AUDET, MAXIME; GUERETTE, PAUL ANDRE; WRAY, LINDSAY; SUBLER, NICOLE ELIZABETH; ZULFIQAR, ZOYA NASIR; LI, HUA; WANG, JESSICA
To: BOLT THREADS, INC.
Reel/Frame 058151/0503 →
Continuity (3)
Continuation 16141787 · Sep 25, 2018
Provisional Application 62563022 · Sep 25, 2017
Related Publication 20220127756A1 · Apr 28, 2022
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