Dipeptide Comprising a Non-Proteogenic Amino Acid

Abstract

Described is a dipeptide comprising a non-proteogenic amino acid, methods of making such and methods of using said dipeptide in a process of making a polypeptide or protein comprising one or more non-proteogenic amino acids.

1 . A dipeptide of Chem. 1: 
       
         
           
           
               
               
           
         
         wherein 
         R1 is H or an amino protecting group, and R2 is an amino protecting group; or
 R1 is a removable alkyl group, and R2 is H or a removable alkyl group; or 
 R1 and R2 are jointly forming a ring; 
 
         R3 is H, or a secondary ammonium cation, a tertiary ammonium cation or a metal cation forming a salt with the carboxylate group; and 
         R4 is absent or an acidic salt. 
       
     
     
         2 . The dipeptide of  claim 1 , wherein the amino protecting group is selected from the group consisting of: tert-Butoxycarbonyl (Boc), Triphenylmethyl (Trt), 2-(p-biphenylyl)-2-propyloxycarbonyl (Bpoc), 9-fluorenylmethyloxycarbonyl (Fmoc), 2-(4-Nitrophenyl)sulfonylethoxycarbonyl (Nsc), Benzyloxycarbonyl (Cbz), Allyloxycarbonyl (Alloc), o-Nitrobenzenesulfonyl (oNBS), p-Nitrobenzenesulfonyl (pNBS), 2,4-Dinitrobenzenesulfonyl (dNBS), 1-(4,4-Dimethyl-2,6-dioxocyclo-Hexylidene)-3-methylbutyl (ivDde) and o- or p-Nitrophenylsulfenyl (Nps). 
     
     
         3 . The dipeptide of  claim 1 , wherein the removable alkyl group is selected from the group consisting of: Benzyl and tert-Butyl. 
     
     
         4 . The dipeptide of  claim 1 , wherein, when R1 and R2 are jointly forming a ring, the jointly formed ring is selected from the group consisting of: Phatalimide and 1,3,5-dioxazine. 
     
     
         5 . The dipeptide of  claim 1 , wherein R1 is H or an amino protecting group selected from the group consisting of: Boc, Trt, Bpoc, Fmoc, Nsc, Cbz, Alloc, oNBS, pNBS, dNBS, ivDde and Nps, and R2 is an amino protecting group selected from the group consisting of: Boc, Trt, Bpoc, Fmoc, Nsc, Cbz, Alloc, oNBS, pNBS, dNBS, ivDde and Nps; or
 R1 is a removable alkyl group selected from the group consisting of: Benzyl and tert-Butyl, and R2 is H or a removable alkyl group selected from the group consisting of: Benzyl and tert-Butyl; or   R1 and R2 are jointly forming a ring selected from the group consisting of: Phatalimide and 1,3,5-dioxazine;   R3 is H, or a secondary ammonium cation, a tertiary ammonium cation, an alkali metal cation or an alkaline earth metal cation forming a salt with the carboxylate group; and   R4 is absent or an acidic salt selected from the group consisting of: A salt of TFA, a salt of HCl, a salt of HBr and a salt of hydrogensulfate.   
     
     
         6 . The dipeptide of  claim 1 , wherein R4 is absent. 
     
     
         7 . The dipeptide of  claim 1 , wherein R4 is an acidic salt selected from the group consisting of: a salt of TFA, a salt of HCl, a salt of HBr and a salt of hydrogensulfate. 
     
     
         8 . The dipeptide according to  claim 1 , which is Fmoc-His-Aib-OH of Chem. 2 
       
         
           
           
               
               
           
         
         wherein His is histidine, Aib is the artificial amino acid 2-aminoisobutyric acid, Fmoc is the protection group 9-fluorenylmethyloxycarbonyl and R4 is absent or an acidic salt such as TFA, HCl, HBr or HOAc. 
       
     
     
         9 . The dipeptide according to  claim 1 , which is the TFA salt of Fmoc-His-Aib-OH:
 Fmoc-His-Aib-OH,TFA   wherein His is histidine, Aib is the artificial amino acid 2-aminoisobutyric acid, Fmoc is the protection group 9-fluorenylmethyloxycarbonyl and TFA is trifluoroacetic acid.   
     
     
         10 . The dipeptide according to  claim 1 , which is activated by an activating agent such as a phosphonium based coupling reagent such as (benzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyBOP). 
     
     
         11 . (canceled) 
     
     
         12 . A method for obtaining a polypeptide or protein comprising one or more non-proteogenic amino acids, wherein the method comprises a step of reacting the dipeptide according to  claim 1  with a polypeptide or protein. 
     
     
         13 . A method for obtaining a polypeptide or protein according to  claim 12 , wherein R1 and/or R2 of said dipeptide is removed in a deprotection step under basic conditions. 
     
     
         14 . A method for obtaining a polypeptide or protein according to  claim 12 , wherein pH of the aqueous media is between pH 8.7 and pH 9.4. 
     
     
         15 . The method for obtaining a polypeptide or protein according to  claim 12 , wherein said dipeptide is reacted with the α-N-terminal of the polypeptide or protein.