Chemical Synthesis of Insulin Analogs through a Novel Precursor

Insulin remains a challenging synthetic target due in large part to its two-chain, disulfide-constrained structure. Biomimetic single chain precursors inspired by proinsulin that utilize short peptides to join the A and B chains can dramatically enhance folding efficiency. Systematic chemical analys...

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Bibliographic Details
Published in:ACS chemical biology 2014-03, Vol.9 (3), p.683-691
Main Authors: Zaykov, Alexander N, Mayer, John P, Gelfanov, Vasily M, DiMarchi, Richard D
Format: Article
Language:English
Subjects:
Alanine - chemistry
Alanine - genetics
Amino Acid Sequence
Amino Acid Substitution
HEK293 Cells
Humans
Insulin - analogs & derivatives
Insulin - chemical synthesis
Insulin - chemistry
Insulin - genetics
Insulin-Like Growth Factor I - genetics
Models, Molecular
Molecular Sequence Data
Peptides - chemistry
Proinsulin - chemistry
Protein Folding
Protein Precursors - chemistry
Protein Precursors - genetics
Protein Refolding
Transfection
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Summary:Insulin remains a challenging synthetic target due in large part to its two-chain, disulfide-constrained structure. Biomimetic single chain precursors inspired by proinsulin that utilize short peptides to join the A and B chains can dramatically enhance folding efficiency. Systematic chemical analysis of insulin precursors using an optimized synthetic protocol identified a 49 amino acid peptide named DesDi, which folds with high efficiency by virtue of an optimized structure and could be proteolytically converted to bioactive two-chain insulin. In subsequent applications, we observed that the folding of the DesDi precursor was highly tolerant to amino acid substitution at various insulin residues. The versatility of DesDi as a synthetic insulin precursor was demonstrated through the preparation of several alanine mutants (A10, A16, A18, B12, B15), as well as ValA16, an analog that was unattainable in prior reports. In vitro bioanalysis highlighted the importance of the native, hydrophobic residues at A16 and B15 as part of the core structure of the hormone and revealed the significance of the A18 residue to receptor selectivity. We propose that the DesDi precursor is a versatile synthetic intermediate for the preparation of diverse insulin analogs. It should enable a more comprehensive analysis of function to insulin structure than might not be otherwise possible through conventional approaches.
ISSN:1554-8929
1554-8937
DOI:10.1021/cb400792s