title: Scope and Limitations of Fmoc Chemistry SPPS-Based Approaches to the Total Synthesis of Insulin Lispro via Ester Insulin.
abstract: We have systematically explored three approaches based on 9-fluorenylmethoxycarbonyl (Fmoc) chemistry solid phase peptide synthesis (SPPS) for the total chemical synthesis of the key depsipeptide intermediate for the efficient total chemical synthesis of insulin. The approaches used were: stepwise Fmoc chemistry SPPS; the "hybrid method", in which maximally protected peptide segments made by Fmoc chemistry SPPS are condensed in solution; and, native chemical ligation using peptide-thioester segments generated by Fmoc chemistry SPPS. A key building block in all three approaches was a Glu[O-β-(Thr)] ester-linked dipeptide equipped with a set of orthogonal protecting groups compatible with Fmoc chemistry SPPS. The most effective method for the preparation of the 51 residue ester-linked polypeptide chain of ester insulin was the use of unprotected peptide-thioester segments, prepared from peptide-hydrazides synthesized by Fmoc chemistry SPPS, and condensed by native chemical ligation. High-resolution X-ray crystallography confirmed the disulfide pairings and three-dimensional structure of synthetic insulin lispro prepared from ester insulin lispro by this route. Further optimization of these pilot studies could yield an efficient total chemical synthesis of insulin lispro (Humalog) based on peptide synthesis by Fmoc chemistry SPPS.
authors.affiliation: Department of Chemistry, Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL, 60637, USA.
authors.forename: Balamurugan
authors.initials: B
authors.lastname: Dhayalan
authors.affiliation: Department of Chemistry, Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL, 60637, USA.
authors.forename: Kalyaneswar
authors.initials: K
authors.lastname: Mandal
authors.affiliation: Department of Biochemistry, Case Western Reserve University, Cleveland, OH, 44106, USA.
authors.forename: Nischay
authors.initials: N
authors.lastname: Rege
authors.affiliation: Department of Biochemistry, Case Western Reserve University, Cleveland, OH, 44106, USA.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Weiss
authors.affiliation: Bachem AG, Hauptstrasse 144, 4416, Bubendorf, Switzerland.
authors.forename: Simon H
authors.initials: SH
authors.lastname: Eitel
authors.affiliation: Bachem AG, Hauptstrasse 144, 4416, Bubendorf, Switzerland.
authors.forename: Thomas
authors.initials: T
authors.lastname: Meier
authors.affiliation: Bachem AG, Hauptstrasse 144, 4416, Bubendorf, Switzerland.
authors.forename: Ralph O
authors.initials: RO
authors.lastname: Schoenleber
authors.affiliation: Department of Chemistry, Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL, 60637, USA.
authors.forename: Stephen B H
authors.initials: SB
authors.lastname: Kent
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D004220:Disulfides / Q000737:chemistry; D005449:Fluorenes / Q000737:chemistry; D007004:Hypoglycemic Agents / Q000138:chemical synthesis* / Q000737:chemistry; D061268:Insulin Lispro / Q000138:chemical synthesis* / Q000737:chemistry; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D060327:Solid-Phase Synthesis Techniques
chemical_list: C496789:9-fluorenylmethoxycarbonyl; D004220:Disulfides; D005449:Fluorenes; D007004:Hypoglycemic Agents; D061268:Insulin Lispro
keywords: Fmoc chemistry SPPS; hybrid synthesis; insulin lispro; native chemical ligation; total synthesis