Back to results
US 6,307,018 B1Granted
General chemical ligation
AssigneeThe Scripps Research Institute
InventorsStephen B. H. Kent, Tom W. Muir, Philip E. Dawson, Steven J. Bark, Lynne Canne Bannen
Life of this patent
Estimating the term…
Abstract
Disclosed herein is a process whereby oligopeptides are ligated to form ligation peptide products. In the first step, two starting oligopeptides are ligated to form an intermediate having an amino-thioester linkage. In the second step, the aminothioester linkage undergoes a rearrangement to form a peptide having an N-substituted amide linkage. In an optional third step, the N-substitution of the amide linkage is chemically removed to form a native peptide linkage.
Claims8
A method for ligating a first oligopeptide with a second oligopeptide end-to-end to produce an oligopeptide product, the method comprising the following steps:
- Step A:
- admixing the first and second oligopeptides in a reaction solution including a thiol that is present in a catalytic amount, the first oligopeptide including a C-terminal residue having a C-termninal thioester, the second oligopeptide including an N-terminal amino acid residue having a secondary amino group, the secondary amino group being attached to an N-linked auxiliary functional group having an unoxidized sulfhydryl moiety, the C-terminal residue being glycine when the N- terminal residue is non-glycine, the N-terminal residue being glycine when the C-terminal residue is non-glycine;
- Step B:
- condensing the unoxidized sulfhydryl moiety of the N-linked auxiliary functional group on the first oligopeptide with the C-terminal thioester of the second oligopeptide for a time and under conditions effective to produce an intermediate oligopeptide which spontaneously rearranges to form the oligopeptide product,
- wherein said intermediate oligopeptide consists of the first oligopeptide and the second oligopeptide bonded together via an aminothioester bond, and said oligopeptide product consists of the first oligopeptide and the second oligopeptide bonded together via an amide bond, said amide bond bearing the N-linked auxiliary functional group;
- and
- Step C:
- isolating the oligopeptide product having an amide bond that bears the N-linked auxiliary functional group.
5 more elements
A method for ligating a first oligopeptide with a second oligopeptide end-to-end to produce an oligopeptide product, the method comprising the following steps:
- Step A:
- admixing the first and second oligopeptides in a reaction solution including a thiol that is present in a catalytic amount, the first oligopeptide including a C-terminal residue having a C-terminal thioester, the second oligopeptide including an N-terminal amino acid residue having a secondary amino group, the secondary amino group being attached to an N-linked auxiliary functional group having an unoxidized sulfhydryl moiety, the C-terminal residue being glycine when the N- terminal residue is non-glycine, the N-terminal residue being glycine when the C-terminal residue is non-glycine;
- Step B:
- condensing the unoxidized sulfhydryl moiety of the N-linked auxiliary functional group on the first oligopeptide with the C-terminal thioester of the second oligopeptide for a time and under conditions effective to produce an intermediate oligopeptide which spontaneously rearranges to form the oligopeptide product;
- wherein said intermediate oligopeptide consists of the first oligopeptide and the second oligopeptide bonded together via an aminothioester bond, and said oligopeptide product consists of the first oligopeptide and the second oligopeptide bonded together via an amide bond, said amide bond bearing the N-linked auxiliary functional group;
- Step C:
- reacting the oligopeptide product of step B with a reducing agent for a time and under conditions effective to remove the N-linked auxiliary functional group that is present on said amide, thereby producing a native peptide bond;
- and
- Step D:
- isolating the oligopeptide product having the native peptide bond.
7 more elements
A method for ligating a first oligopeptide with a second oligopeptide end-to-end to produce an oligopeptide product, the method comprising the following steps:
- Step A:
- admixing the first and second oligopeptides in a reaction solution including a thiol that is present in a catalytic amount, the first oligopeptide including a C-terminal residue having a C-terminal thioester, the second oligopeptide including an N-terminal amino acid residue bearing an N-linked auxiliary functional group having an unoxidized sulfhydryl moiety and being respresented by —O—(CH2)n—SH, wherein 1≦n≦2, the C-termninal residue being glycine when the N-terminal residue is non-glycine, the N-terminal residue being glycine when the C-terminal residue is non-glycine;
- Step B:
- condensing the unoxidized sulfhydryl moiety of the N-linked auxiliary functional group on the first oligopeptide with the C-terminal thioester of the second oligopeptide for a time and under conditions effective to produce an intermediate oligopeptide which spontaneously rearranges to form the oligopeptide product;
- wherein said intermediate oligopeptide consists of the first oligopeptide and the second oligopeptide bonded together via an aminothioester bond, and said oligopeptide product consists of the first oligopeptide and the second oligopeptide bonded together via an amide bond, said amide bond bearing the N-linked auxiliary functional group;
- and
- Step C:
- isolating the oligopeptide product having an amide bond that bears the N-linked auxiliary functional group.
5 more elements
A method for ligating a first oligopeptide with a second oligopeptide end-to-end to produce an oligopeptide product, the method comprising the following steps:
- Step A:
- admixing the first and second oligopeptides in a reaction solution including a thiol that is present in a catalytic amount, the first oligopeptide including a C-terminal residue having a C-terminal thioester, the second oligopeptide including an N-terminal amino acid residue bearing an N-linked auxiliary functional group having an unoxidized sulfhydryl moiety and being respresented by —O—(CH2)n—SH, wherein 1≦n≦2, the C-terminal residue being glycine when the N-terminal residue is non-glycine, the N-terminal residue being glycine when the C-terminal residue is non-glycine;
- Step B:
- condensing the unoxidized sulfhydryl moiety of the N-linked auxiliary functional group on the first oligopeptide with the C-terminal thioester of the second oligopeptide for a time and under conditions effective to produce an intermediate oligopeptide which spontaneously rearranges to form the oligopeptide product;
- wherein said intermediate oligopeptide consists of the first oligopeptide and the second oligopeptide bonded together via an aminothioester bond, and said oligopeptide product consists of the first oligopeptide and the second oligopeptide bonded together via an amide bond, said amide bond bearing the N-linked auxiliary functional group;
- Step C:
- reacting the oligopeptide product of step B with a reducing agent for a time and under conditions effective to remove the N-linked auxiliary functional group that is present on said amide, thereby producing a native peptide bond;
- and
- Step D:
- isolating the oligopeptide product having the native peptide bond.
7 more elements
Dependencies are inferred from the claim text.