The α-Ketoacid-Hydroxylamine (KAHA) Amide-Forming Ligation is a chemical reaction that is used to join two unprotected fragments in

peptide synthesis In organic chemistry, peptide synthesis is the production of peptides, compounds where multiple amino acids are linked via amide bonds, also known as peptide bonds. Peptides are chemically synthesized by the condensation reaction of the carboxyl ...

.{{cite journal, last1=Pusterla, first1=Ivano, last2=Bode, first2=Jeffrey W., title=The Mechanism of the α-Ketoacid-Hydroxylamine Amide-Forming Ligation, journal=Angewandte Chemie, volume=124, issue=2, year=2012, pages=528–531, issn=0044-8249, doi=10.1002/ange.201107198 It is an alternative to the Native Chemical Ligation (NCL). KAHA Ligation was developed by Jeffrey W. Bode group at

ETH Zürich (colloquially) , former_name = eidgenössische polytechnische Schule , image = ETHZ.JPG , image_size = , established = , type = Public , budget = CHF 1.896 billion (2021) , rector = Günther Dissertori , president = Joël Mesot , a ...

(previously University of Pennsylvania).

Overview

An α-ketoacid at the C-terminus of one peptide fragment reacts with a

hydroxylamine Hydroxylamine is an inorganic compound with the formula . The material is a white crystalline, hygroscopic compound.Greenwood and Earnshaw. ''Chemistry of the Elements.'' 2nd Edition. Reed Educational and Professional Publishing Ltd. pp. 431–43 ...

at the N-terminus of another to form a peptide bond (amide bond). The reaction can happen in the presence of unprotected side chains, and it does not require any coupling reagents or

catalyst Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...

s. The only byproducts are water and CO₂. KAHA Ligation Type I

The first reported protein synthesized by KAHA ligation was human

GLP-1 Glucagon-like peptide-1 (GLP-1) is a 30- or 31-amino-acid-long peptide hormone deriving from the tissue-specific posttranslational processing of the proglucagon peptide. It is produced and secreted by intestinal enteroendocrine L-cells and certa ...

(7-36). Since then, a variety of small proteins (up to 200 residues) have been synthesized, including

ubiquitin Ubiquitin is a small (8.6 kDa) regulatory protein found in most tissues of eukaryotic organisms, i.e., it is found ''ubiquitously''. It was discovered in 1975 by Gideon Goldstein and further characterized throughout the late 1970s and 1980s. Fo ...

and other similar modifier proteins, hormone proteins, nitrophorin 4, S100A4 and cyclic proteins. C-terminal ketoacid monomers are pre-loaded on resin via a linker for Fmoc-SPPS (Fmoc-based

solid phase peptide synthesis In organic chemistry, peptide synthesis is the production of peptides, compounds where multiple amino acids are linked via amide bonds, also known as peptide bonds. Peptides are chemically synthesized by the condensation reaction of the carboxy ...

). Initial research utilised sulfur ylide linkers, but more recently the group developed acid- and photo-labile ketoacid monomers that can be loaded directly on Rink Amide resin. The most commonly used N-terminal hydroxylamine is the 5-oxaproline, which results in a

homoserine Homoserine (also called isothreonine) is an α-amino acid with the chemical formula HO2CCH(NH2)CH2CH2OH. -Homoserine is not one of the common amino acids encoded by DNA. It differs from the proteinogenic amino acid serine by insertion of an additi ...

residue after ligation and O-N rearrangement.

References

Peptides