MOENOMYCIN BIOSYNTHESIS-RELATED COMPOSITIONS AND METHODS OF USE THEREOF

• US 20100279980A1
• Filed: 08/13/2007
• Published: 11/04/2010
• Est. Priority Date: 08/11/2006
• Status: Active Grant

• Current Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Original Assignee: President and Fellows of Harvard College (Harvard Corporation)
• Inventors: Walker, Suzanne, Ostash, Bohdan Omelyanovich
• Granted Patent: US 9,115,358 B2
• Time in Patent Office: Days
• US Class Current: 514/54
• CPC Class Codes: A61K 31/7028 Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
• CPC Class Codes: C12N 15/52 Genes encoding for enzymes or proenzymes
• CPC Class Codes: C12P 19/44 Preparation of O-glycosides, e.g. glucosides polysaccharides and not substituted disaccharides C12P19/04, C12P19/12

Abstract

The methods and compositions described herein relate to the identification, isolation, and characterization of genes which encode proteins useful for the biosynthesis of transglycosylase inhibitors such as moes. The methods and compositions also relate to the production of such proteins, and their use in the synthesis of moes, the expression of moes, and the production of modified moes.

Claims

2. A nucleic acid composition comprising a one or more sequences selected from the group consisting of:

SEQ ID NOs;

3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, and 25, fragments thereof, or natural or artificial variants thereof.

3. The composition of claim 2, wherein the nucleic acid composition comprises an open reading frame.

4. The composition of claim 3, wherein the nucleic acid composition is in an expression vector.

5. The composition of claim 4, wherein the expression vector is expressed in a bacterial, insect, yeast or mammalian cell.

7. A composition comprising a one or more polypeptides selected from the group consisting of:

moeA4, moeB4, moeC4, moeB5, moe A5, moeD5, moeJ5, moeE5, moeF5, moeH5, moeK5, moeM5, moeN5, moeO5, moeX5, moeP5, moeR5, moeS5, moeGT1, moeGT2, moeGT3, moeGT4, and moeGT5, fragments thereof, or a natural or artificial variants thereof.

1. The composition of claim 7, wherein the one or more polypeptides is encoded by a nucleic acid sequence selected from the group consisting of:

SEQ ID NOs;

3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, and 25, fragments, or natural or artificial variants thereof.

6. The composition of claim 7, wherein the one or more polypeptides comprises a sequence selected from the group consisting of:

SEQ ID NOs;

26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, and 48, natural or artificial variants, or fragments thereof.

8. (canceled)

9. A moenomycin A molecule, a moenomycin derivative or a moenomycin intermediate produced by a Streptomyces strain carrying a one or more mutant or inactivated genes, wherein the one or more mutant or inactivated genes is selected from the group consisting of:

moeA4, moeB4, moeC4, moeB5, moe A5, moeD5, moeJ5, moeE5, moeF5, moeH5, moeK5, moeM5, moeN5, moeO5, moeX5, moeP5, moeR5, moeS5, moeGT1, moeGT2, moeGT3, moeGT4, and moeGT5.

10. The moenomycin A molecule of claim 9, wherein the Streptomyces strain is Streptomyces ghanaensis ATCC14627 or Streptomyces lividans TK24.

11. (canceled)

12. A method of synthesizing or modifying a moenomycin, a moenomycin derivative, or a moenomycin intermediate wholly or partially in vitro comprising:

reacting a one or more moenomycin precursor, derivative and/or moenomycin intermediate with a one or more polypeptide selected from the group consisting of;

moeA4, moeB4, moeC4, moeB5, moe A5, moeD5, moeJ5, moeE5, moeF5, moeH5, moeK5, moeM5, moeN5, moeO5, moeX5, moeP5, moeR5, moeS5, moeGT1, moeGT2, moeGT3, moeGT4, and moeGT5, under conditions wherein the moenomycin, the moenomycin derivative, or the intermediate is wholly or partially synthesized.

14. A method of treating a subject comprising:

administering a therapeutically effective amount of the moenomycin of claim 12 to the subject.

15. The method of claim 14, wherein the subject is a human.

55. The method according to claim 12, wherein the method further comprises reacting the moenomycin, moenomycin derivative and/or moenomycin intermediate with a one or more reactants selected from the group consisting of:

UDP-sugars, prenyl-pyrophosphates, phosphoglycerate, amino acids, cabamoyl phosphate, ATP and biological cofactors.

13. (canceled)

16. A moenomycin derivative, pharmaceutically acceptable salt, tautomer, and/or ester thereof, wherein the moenomycin derivative has the structure:

17. The moenomycin derivative of claim 16, wherein R and R¹ independently are —

NH₂.

38. The moenomycin derivative, pharmaceutically acceptable salt, tautomer, and/or ester thereof of claim 16, wherein the moenomycin derivative structure is:

43. The moenomycin derivative of claim 38, wherein R⁴ is hydroxyl and R⁶ is

44. A pharmaceutical composition comprising the moenomycin derivative of claim 16 and a pharmaceutically acceptable carrier.

18. through 37 (canceled)

39. through 41. (canceled)

42. The moenomycin derivative of claim 41, wherein substituted cycloalkyl is

45. A moenomycin derivative, pharmaceutically acceptable salt, tautomer, and/or ester thereof, wherein the moenomycin derivative has the structure:

49. A pharmaceutical composition comprising the moenomycin derivative of claim 45 and a pharmaceutically acceptable carrier.

46. The moenomycin derivative of claim 45, wherein R⁷ and R⁸ independently are —

NH₂. cm 47 and 48. (canceled)

50. An isolated Streptomyces strain selected from the group consisting of:

Streptomyces ghanaensis, Streptomyces ederensis, Streptomyces geysiriensis, and Streptomyces bambergiensis strain which carries a one or more mutant or inactivated genes, wherein the mutant or inactivated genes are selected from the group consisting of;

moeA4, moeB4, moeC4, moeB5, moe A5, moeD5, moeJ5, moeE5, moeF5, moeH5, moeK5, moeM5, moeN5, moeO5, moeX5, moeP5, moeR5, moeS5, moeGT1, moeGT2, moeGT3, moeGT4, and moeGT5.

51. The isolated Streptomyces strain of claim 50, wherein the Streptomyces ghanaensis strain is Streptomyces ghanaensis ATCC14627.

52. An isolated recombinant cell expressing a one or more polypeptides, or fragments, or natural, or artificial variants thereof selected from the group consisting of:

moeA4, moeB4, moeC4, moeB5, moeA5, moeD5, moeJ5, moeE5, moeF5, moeH5, moeK5, moeM5, moeN5, moeO5, moeX5, moeP5, moeR5, moeS5, moeGT1, moeGT2, moeGT3, moeGT4, and moeGT5.

53. The isolated recombinant cell of claim 52, wherein the cell is selected from the group consisting of:

Streptomyces lividans TK24, E. coli, mammalian cells, yeast and insect cells.

54. A moenomycin A molecule derivative or intermediate produced by the isolated recombinant cell of claim 52.

56. (canceled)