Spider silk protein

• US 5,728,810 A
• Filed: 04/19/1995
• Issued: 03/17/1998
• Est. Priority Date: 04/20/1990
• Status: Expired due to Term

• Current Assignee: University of Wyoming
• Original Assignee: University of Wyoming
• Inventors: Xu, Ming, Lewis, Randolph V., Hinman, Michael B.
• Primary Examiner(s): Wax, Robert A.
• Assistant Examiner(s): Lau, Kawai
• Application Number:
  US08/425,069
• Time in Patent Office: 1,063 Days
• Field of Search: 530/353, 530/350
• US Class Current: 530/353
• CPC Class Codes: C07K 14/195 from bacteria
• CPC Class Codes: C07K 14/43518 from spiders
• CPC Class Codes: C07K 2319/00 Fusion polypeptide

Abstract

The present invention is directed to isolated cDNA which codes for spider silk protein or a fragment or variant thereof, a replicable vector containing cDNA which codes for spider silk protein and which is capable of expressing spider silk protein, a transformed cell or microorganism containing cDNA which codes for spider silk protein or a fragment thereof which is capable of expressing spider silk protein and products, such as fibers, which may be manufactured utilizing the recombinant protein of the present invention.

Claims

1. A purified recombinant spider silk protein, having a molecular weight of at least 16,000 daltons, comprising a polypeptide selected from the group consisting of:

a polypeptide having the amino acid sequence of SEQ. ID. NO.;

2;

a polypeptide having the amino acid sequence of SEQ. ID. NO.;

4;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

5;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

6;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

7;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

12 linked by a peptide bond to the amino terminus of the amino acid sequence of SEQ. ID. NO.;

13;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

8 linked by a peptide bond to the amino terminus of the amino acid sequence of SEQ. ID. NO.;

12 in turn linked by a peptide bond to the amino acid sequence of SEQ. ID. NO.;

9;

a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.;

8 linked by a peptide bond to the amino terminus of the amino acid sequence of SEQ. ID. NO.;

12 in turn linked by a peptide bond to the amino acid sequence of SEQ. ID. NO.;

10.

2. The polypeptide of claim 1 that is a polypeptide having the amino acid sequence of SEQ. ID. NO.:

2.

3. The polypeptide of claim 1 that is a polypeptide having the amino acid sequence of SEQ. ID. NO.:

4.

4. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.:

5.

5. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.:

6.

6. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.:

7.

7. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.:

12 linked by a peptide bond to the amino terminus of the amino acid sequence of SEQ. ID. NO.;

13.

8. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ. ID. NO.:

8 linked by a peptide bond to the amino terminus of the amino acid sequence of SEQ. ID. NO.;

12 in turn linked by a peptide bond to the amino acid sequence of SEQ. ID. NO.;

9.

9. The polypeptide of claim 1 that is a polypeptide comprising tandem repeats of the amino acid sequence of SEQ ID. NO.:

8 linked by a peptide bond to the amino acid sequence of SEQ. ID. NO.;

12 in turn linked by a peptide bond to the amino acid sequence of SEQ. ID. NO.;

10.

10. A purified silk protein which comprises a polypeptide having an amino acid sequence comprising repeating units of the hexamer GGXGZG, SEQ. ID. NO.:

69, wherein X and Z are any amino acid, but are most often glutamine or alanine.

11. The silk protein of claim 10, wherein X and Z are glutamine or alanine.

12. The silk protein of claim 10, wherein the repeat units are separated by varying numbers of alanine or serine residues.

13. The silk protein of claim 10, which further comprises repeating units having the sequence (A)_m GGAGQGGYGGLGGQG, wherein m is 6 or 7, SEQ. ID. NO.:

5.

14. The silk protein of claim 10, which further comprises repeat units having the amino acid sequence AGRGGXGGZGAG(A)mGGAGQGGBGGLGGQG, where X, Z and B are leucine, tyrosine or glutamine and X is not the same amino acid as Z and wherein m is 6 or 7, SEQ. ID. NO.:

7.

15. The protein of claim 10, which further comprises nucleotides encoding repeat units having the amino acid sequence AGRGGXGGZGAG(A)_m GGAGQGGBGGLGGQG, where X, Z and B are leucine, tyrosine or glutamine and X is not the same amino acid as Z, SEQ. ID. NO.:

7, and wherein m is 6 or 7.

16. A purified silk protein which comprises a polypeptide having an amino acid sequence comprising repeating units of the hexamer GGXGZG, SEQ. ID. NO.:

69, and of polyalanine, wherein X and Z can be any amino acid, but wherein about 46% of X residues are alanine and about 49% of X residues are glutamine and wherein about 49% of Z residues are alanine and about 46% of Z residues are glutamine and wherein the number of alanine residues in said polyalanine ranges from 4 to 7.

17. A purified silk protein which comprises a polypeptide having an amino acid sequence comprising repeating units having the amino acid sequence GPGQQGPGYYGPGQQGPSGPGS, SEQ. ID. NO.:

8.

18. A purified protein which comprises repeating units which contain a sequence represented by the formula

View Dependent Claims (23, 24)

23. A protein of any one of claims 18, 19, 20 and 21, wherein said protein has a molecular weight of 16,000 to 300,000 daltons.

24. A purified protein according to claim 18, wherein for each repeat, m and n are both 10.

19. A purified silk protein which comprises repeats of a polypeptide that is described by the general formula

20. A purified silk protein which comprises repeats of a polypeptide that is described by the general formula

21. A purified protein comprising repeating units selected from the group consisting of ##STR13## and mixtures thereof.

22. A purified protein comprising a polypeptide having an amino acid sequence comprising repeating units of the hexamer GGXGZG, wherein X and Z are any amino acid, SEQ. ID. NO.:

69, but are most often glutamine or alanine and further comprising repeating units having the amino acid sequence (A)_m GGAGQGGYGGLGGQG, wherein m is 6 or 7, SEQ. ID. NO.;

5.