Isolated DNA coding for spider silk protein, a replicable vector and a transformed cell containing the DNA

• US 5,989,894 A
• Filed: 10/04/1994
• Issued: 11/23/1999
• 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:
  US08/317,844
• Time in Patent Office: 1,876 Days
• Field of Search: 530/353, 536/23.5, 435/320.1, 435/69.1, 435/252.3, 435/252.33
• US Class Current: 435/252.3
• 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. An isolated DNA which codes for a protein comprising the amino acid sequence described in SEQ. ID. NO. 2.

12. A replicable vector containing the DNA of claim 1,2,3,4,5,6 or 9.

13. A host cell comprising the vector of claim 12.

14. A replicable vector according to claim 12, wherein said protein is one having a molecular weight from 100,000 to 300,000 daltons.

2. An isolated DNA which codes for a protein comprising the amino acid sequence described in SEQ. ID. NO. 4.

3. An isolated DNA which encodes a protein comprising repeating units having amino acid sequences selected from the group consisting of SEQ ID NO:

26, SEQ ID NO;

27, SEQ ID NO;

28, SEQ ID NO;

29, SEQ ID NO;

30, SEQ ID NO;

31, SEQ ID NO;

32, SEQ ID NO;

33, SEQ ID NO;

34, SEQ ID NO;

35, SEQ ID NO;

36, and SEQ ID NO;

37, and mixtures thereof.

4. An isolated DNA molecule encoding a protein which comprises nucleotides encoding repeat units having the amino acid sequence Ala-Gly-Arg-Gly-Gly-Xaa-Gly-Gly-Zaa-Gly-Ala-Gly- (Ala)_m -Gly-Gly-Ala-Gly-Glu-Gly-Gly-Baa-Gly-Gly-Leu-Gly-Gly-Glu-Gly (SEO ID NO:

7), wherein m is 6 or 7 and wherein Xaa, Zaa and Baa are leucine, tyrosine or glutamine and Xaa is not the same amino acid as Zaa.

15. An isolated DNA molecule according to claim 4, wherein Xaa is glutamine.

16. An isolated DNA molecule according to claim 4, wherein the repeat units are separated by varying numbers of alanine or serine residues.

5. An isolated DNA which encodes a protein comprising repeating units having amino acid sequences selected from the group consisting of SEQ ID NO:

38, SEQ ID NO;

39, SEQ ID NO;

40, SEQ ID NO;

41, SEQ ID NO;

42, SEQ ID NO;

43, SEQ ID NO;

44, SEQ ID NO;

45, SEQ ID NO;

46, SEQ ID NO;

47, SEQ ID NO;

48, SEQ ID NO;

50, SEQ ID NO;

51, SEQ ID NO;

52, SEQ ID NO;

53, SEQ ID NO;

55, SEQ ID NO;

56, SEQ ID NO;

58, SEQ ID NO;

59, SEQ ID NO;

60, SEQ ID NO;

61, and SEQ ID NO;

62, and mixtures thereof.

6. An isolated cDNA which codes for a protein comprising repeating units wherein said repeating units are comprised of a sequence represented by the formula

View Dependent Claims (7, 8, 9, 10, 11, 17, 18)

7. The cDNA of claim 6, wherein p is 15 to 50.

8. The cDNA of claim 6, wherein p is 25 to 100.

9. An isolated cDNA according to claim 6, wherein m is 4 to 10, n is 10 to 20, and p is an integer of 1 to 100 and each X, which may be the same or different, is selected from the group consisting of G, A, Q, Y and L, wherein at least 50% of the X'"'"'s are G.

10. The cDNA of claim 9, wherein p is 15 to 50.

11. The cDNA of claim 9, wherein p is 25 to 100.

17. An Escherichia coli host cell comprising a plasmid comprising the cDNA of claim 8.

18. An Escherichia coli host cell comprising a plasmid comprising the cDNA of claim 9.

19. A plasmid pLM-4 which is contained in ATCC Deposit No. 68567.

20. A plasmid pMB-2 which is contained in ATCC Deposit No. 68568.