Claims
- 1. A method for detecting and/or characterizing binding between an isolated oligosaccharide and a binding partner, comprising the steps of:
contacting an isolated oligosaccharide with a binding partner under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; and detecting the presence of the oligosaccharide-binding partner complex.
- 2. A method for determining the requirement of a binding partner for a functional group on an oligosaccharide in order for the binding partner to bind to the oligosaccharide, the method comprising the steps of:
contacting a modified oligosaccharide comprising at least one functional group with a binding partner under conditions that allow binding of the modified oligosaccharide with the binding partner to form a modified oligosaccharide-binding partner complex; and detecting the presence of the modified oligosaccharide-binding partner complex, wherein the presence of the modified oligosaccharide-binding partner complex is indicative of a requirement for the functional group on the oligosaccharide for binding to the binding partner.
- 3. A method for identifying an agent capable of altering the binding of an oligosaccharide with a binding partner, the method comprising the steps of:
contacting an oligosaccharide with a binding partner under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; contacting the oligosaccharide with a binding partner and an agent under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; and detecting the presence of the oligosaccharide-binding partner complex, wherein a difference in the amount of the oligosaccharide-binding partner complexes in the presence of the agent compared to the amount of the oligosaccharide-binding partner complexes in the absence of the agent is indicative that the agent inhibits or enhanced the binding of the oligosaccharide with the binding partner.
- 4. A method for determining the binding affinity for an oligosaccharide with a binding partner, comprising:
contacting an oligosaccharide with various quantities of a binding partner under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; detecting the presence and/or quantity of the oligosaccharide-binding partner complexes for each quantity of binding partner; and calculating a dissociation constant for the oligosaccharide-binding partner complex.
- 5. A method for determining the minimal binding length of an oligosaccharide for a binding partner, comprising:
contacting samples of oligosaccharides of varying lengths with samples of a binding partner under conditions that allow binding of the oligosaccharides of varying lengths with the binding partner to form oligosaccharide-binding partner complexes; and detecting the presence of the oligosaccharide-binding partner complexes, wherein detection of an oligosaccharide-binding partner complex comprising the shortest oligosaccharide is indicative of the minimum binding length of the oligosaccharide for binding the binding partner.
- 6. A method for identifying and/or characterizing a binding partner that binds to an oligosaccharide, comprising:
contacting an oligosaccharide with a candidate binding partner or a heterogenous mixture containing a candidate binding partner under conditions that allow binding of the oligosaccharide with the candidate binding partner to form an oligosaccharide-candidate binding partner complex; and detecting the presence of the oligosaccharide-candidate binding partner complex.
- 7. A method of screening a library of oligosaccharides for binding to a candidate binding partner, the method comprising:
contacting an oligosaccharide library comprising a plurality of oligosaccharides, which oligosaccharides comprise at least one functional group, with a candidate binding partner or a heterogenous mixture containing the candidate binding partner under conditions that allow binding of the oligosaccharides with the candidate binding partner to form an oligosaccharide-candidate binding partner complex; and detecting the presence of the oligosaccharide-candidate binding partner complex.
- 8. The method of any of claims 1-7, further comprising the step of modifying the oligosaccharide to contain at least one functional group.
- 9. The method of claim 1, wherein the oligosaccharide comprises between 4 and 300 saccharides.
- 10. The method of claim 1, wherein the oligosaccharide comprises at least a pentasaccharide.
- 11. The method of claim 1, wherein the oligosaccharide comprises heparan sulfate.
- 12. The method of claim 1, wherein the oligosaccharide comprises heparan sulfate, chondroitin sulfate, dermatan sulfate, and/or keratan sulfate.
- 13. The method of claim 1, wherein the oligosaccharide is manufactured in vitro.
- 14. The method of claim 1, wherein the oligosaccharide is derived from an in vivo tissue sample.
- 15. The method of claim 1, wherein the oligosaccharide is modified in vitro.
- 16. The method of claim 1, wherein the oligosaccharide is modified in vitro with 3-OST-1 and/or 6-OST-1.
- 17. The method of claim 1, wherein the oligosaccharide is modified in vitro with at least one sulfotransferase selected from the group consisting of 2-OST, 3-OST-2, 3-OST-3A, 3-OST-4, 6-OST-2A, 6-OST-2B, and 6-OST-3.
- 18. The method of claim 1, wherein the oligosaccharide is modified in vitro with an N-deacetylase/N-sulfotransferase (NDST).
- 19. The method of claim 1, wherein the NDST is selected from the group consisting of NDST1, NDST2, NDST3, and NDST4.
- 20. The method of claim 1, wherein the binding partner is a protein.
- 21. The method of claim 1, wherein the binding partner is selected from the group consisting of antithrombin III (AT-III), a fibroblast growth factor (FGF), fibroblast growth factor receptor (FGFR), vascular endothelial growth factor (VEGF), placental growth factor (PlGF), heparin-bindig EGF-like growth factor, hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-beta), platelet-derived growth factor (PDGF), pleiotrophin, platelet factor-4 (PF-4), interleukin-8 (IL-8), macrophage inflammatory protein-1 (MIP-1), interferon-g-inducible protein-10 (IP-10), interferon-gamma (IFN-gamma), and HIV-Tat transactivating factor.
- 22. The method of claim 1, wherein the binding partner is a peptide.
- 23. The method of claim 1, wherein the binding partner is an antibody.
- 24. The method of claim 1, wherein the binding partner comprises a detectable label.
- 25. The method of claim 1, wherein the oligosaccharide comprises a detectable label.
- 26. The method of claim 24 or 25, wherein the detectable label is a radioactive label.
- 27. The method of claim 24 or 25, wherein the detectable label is a non-radioactive label selected from the group consisting of biotin, fluorescein, and green fluorescent protein.
- 28. The method of claim 3, wherein the agent blocks infection of a pathogen.
- 29. The method of claim 28, wherein the pathogen is a virus, bacteria, prion, fungus, yeast, or parasite.
- 30. The method of claim 3, wherein the agent is a pro-coagulant.
- 31. The method of claim 3, wherein the agent inhibits cell growth.
- 32. The method of claim 3, wherein the agent inhibits cell adhesion.
- 33. The method of claim 3, wherein the agent inhibits inflammation.
- 34. The method of claim 3, wherein the agent inhibits enzyme activity.
- 35. The method of claim 3, wherein the agent is an agonist to oligosaccharide-binding partner binding.
- 36. The method of claim 1, wherein said detecting step comprises subjecting the oligosaccharide-binding partner complex to migration through a medium.
- 37. The method of claim 2, wherein said detecting step comprises subjecting the modified oligosaccharide-binding partner complex to migration through a medium.
- 38. The method of claim 36 or 37, wherein the medium is a gel.
- 39. The method of claim 38, wherein the medium is a gel selected from the group consisting of a vertical gel, a horizontal gel, and a capillary gel.
- 40. The method of claim 1, wherein the detecting step comprises gel electrophoresis.
- 41. The method of claim 40, wherein the gel is subjected to autoradiography.
- 42. The method of claim 1, wherein said detecting step comprises a gel mobility shift assay.
- 43. An agent identified using the method of claim 3.
- 44. A method of treating a condition associated with altered oligosaccharide-binding partner binding, the method comprising administering an effective amount of an agent identified and/or characterized in claim 3.
- 45. A method for identifying and/or characterizing an enzyme that modifies an oligosaccharide, comprising:
contacting an oligosaccharide with a candidate enzyme or a heterogenous mixture comprising a candidate enzyme under conditions that allow the candidate enzyme to modify the oligosaccharide; contacting the modified oligosaccharide with a binding partner under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; and detecting the presence of the oligosaccharide-binding partner complex, wherein the presence of the oligosaccharide-binding partner complex is indicative of the presence of an enzyme that is capable of modifying the oligosaccharide so that it binds to the binding partner.
- 46. A kit for detecting the binding of an oligosaccharide to a binding partner, comprising:
at least one oligosaccharide, and at least one binding partner that binds to the oligosaccharide.
- 47. The kit of claim 46, wherein the the kit comprises an oligosaccharide library.
- 48. The kit of claim 46, wherein the the kit comprises modifying enzyme.
- 49. The kit of claim 46, wherein the oligosaccharide comprises a detectable label.
- 50. The kit of claim 46, wherein the binding partner comprises a detectable label.
- 51. An isolated composition of matter comprising a 3-O, 6-O sulfated pentasaccharide of heparan sulfate, capable of binding to antithrombin III.
- 52. An isolated composition of matter comprising a 3-O, 6-O sulfated tetrasaccharide of heparan sulfate, capable of binding to fibroblast growth factor 1 (FGF1) and/or fibroblast growth factor receptor 1 (FGFR1).
- 53. A method for diagnosing a condition associated with altered oligoasccharide-binding partner binding in a subject, comprising the steps of:
contacting an oligosaccharide isolated from a subject with a binding partner under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; and comparing the oligosaccharide-binding partner complex with an oligosaccharide-binding partner complex of a normal control, wherein an alteration in the amount or size of the subject's oligosaccharide-binding partner complex compared to the normal control oligosaccharide-binding partner complex is indicative of a condition associated with abnormal oligosaccharide-binding partner binding in the subject.
- 54. A method for diagnosing a condition associated with altered oligoasccharide-binding partner binding in a subject, comprising the steps of:
contacting a binding partner isolated from a subject with an oligosaccharide under conditions that allow binding of the oligosaccharide with the binding partner to form an oligosaccharide-binding partner complex; and comparing the oligosaccharide-binding partner complex with an oligosaccharide-binding partner complex of a normal control, wherein an alteration in the amount or size of the subject's oligosaccharide-binding partner complex compared to the normal control oligosaccharide-binding partner complex is indicative of a condition associated with abnormal oligosaccharide-binding partner binding in the subject.
- 55. The method of claim 53 or 54, wherein the condition is associated with a change in the amount of oligosaccharide and/or binding partner in the subject or a change in oligosaccharide modifications in the subject.
- 56. The method of claim 53 or 54, wherein the condition is associated with a change in the amount or activity of a modifying enzyme that modifies the oligosaccharide.
RELATED APPLICATIONS
[0001] This application claims priority to U.S. Ser. No. 60/326,270, filed Oct. 1, 2001, the entire disclosure of which is hereby incorporated by reference.
Provisional Applications (1)
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Number |
Date |
Country |
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60326270 |
Oct 2001 |
US |