Synthesis of insulin‐like growth factor I using N‐methyl pyrrolidinone as the coupling solvent and trifluoromethane sulphonic acid cleavage from the resin

Abstract

Insulin‐like growth factor I (IGF‐I), a protein of 70 amino acid residues and 3 cystine bridges, has been synthesized by two solid phase Boc methods. The first method used N‐methylpyrrolidinone as the solvent with single coupling cycles while the second synthesis used dimethylformamide and dichloromethane as the solvents with a double‐coupling protocol. In both cases, trifluoroacetic acid/trifluoromethanesulphonic acid cleavage of the peptide from the resin was employed. Purification of the cleavage products followed by removal of the S‐acetamidomethyl protecting groups gave reduced peptides which were then oxidized under conditions favouring the formation of the correct disulphide bonds. The purified synthetic IGF‐I peptides were full agonists of natural IGF‐I in a radioimmunoassay, in an IGF‐I radioreceptor assay, in a bioassay which measures the stimulation of protein synthesis in rat L6 myoblasts and in an IGF‐binding protein competitive binding assay. Moreover, in each of these assays, the synthetic IGF peptides were found to be at least 70% as potent as natural IGF‐I

Chemical synthesis of peptides within the insulin superfamily

“…A subsequent synthesis of IGF‐1 by Wallace et al . reported a yield of ~0.4% . More recently, the preparation of a fluorescently labeled IGF‐2 analog was published by Abell et al .…”

“…The first syntheses of the IGFs were reported by Li and co-workers, who assembled the 70-residue IGF-1 in 1983 with a yield of~1% [102,103] and the 67-residue IGF-2 in 1985 with a yield of~2% [104,105]. A subsequent synthesis of IGF-1 by Wallace et al reported a yield of~0.4% [106,107]. More recently, the preparation of a fluorescently labeled IGF-2 analog was published by Abell et al [108].…”

Improved Fmoc‐based solid‐phase synthesis of homologous peptide fragments of human and mouse prion proteins

“…Although the Boc‐based ones have proven to be successful26, the Fmoc‐based ones have offered varying levels of success26–29. In general, the Fmoc‐based syntheses use polystyrene resins, such as the Wang resin, and differ in their approach in resolving undesired self‐association of the growing sequence, encompassing the use of Hmb‐amide protecting groups26, 27 Dmb‐protected dipeptides29, a mixture of DMF/NMP as solvent30, and/or high temperatures28, 31. However, using extreme conditions for amino acid couplings—especially high temperatures—can decrease the quality of the desired peptide, chiefly due to side reactions such as amino acid racemisation1.…”

Development of the Schedule for Multiple Parallel “Difficult” Peptide Synthesis on Pins

“…The choice of hepatitis C virus envelope protein sites for scanning and the preparation of the peptide list were described elsewhere [7, 9]. The peptide synthesis from Fmoc-amino acids using DIPC as the condensation catalyst, as described in the “Mimotopes” manual and [22], was chosen as a standard procedure, except that 4-methyl-piperidine (4MPIP) was employed instead of piperidine [36, 37] and NMP was used as the Fmoc removal and amino acid attachment solvent instead of DMF [38]. …”