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Exploratory synthesis of peptide–α‐thioester segments spanning the polypeptide sequence of the δ‐opioid receptor, a G protein‐coupled receptor
Author(s) -
Johnson Erik C. B.,
Kent Stephen B. H.
Publication year - 2006
Publication title -
peptide science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/bip.20639
Subject(s) - chemistry , peptide , native chemical ligation , receptor , peptide sequence , combinatorial chemistry , opioid peptide , pentapeptide repeat , transmembrane protein , thioester , chemical ligation , transmembrane domain , g protein coupled receptor , chemical synthesis , biochemistry , stereochemistry , opioid , in vitro , enzyme , gene
We have decided to use the δ‐opioid receptor (372 residues) as a model system to develop methods for the total chemical synthesis of G protein‐coupled receptors. The most important feature of this receptor from a chemical synthesis perspective is the wealth of cysteines spread throughout its sequence, which are required for native chemical ligation. A total of 13 cysteines are located in the the δ‐opioid receptor polypetide chain in both loop and putative transmembrane (TM) regions. We envisioned a synthesis of the polypeptide that would make use of peptide–α‐thioesters ranging from 37 to 63 residues in length. Here, we report data from an exploratory synthesis of such a set of peptide–α‐thioesters. For all seven peptides, the crude material ∼ 30 residues into the synthesis was sufficiently homogeneous to make isolation and purification straightforward. Extension of the peptides to between 40 and 50 residues in length generally produced a significant decrease in the quality of the crude products, although in most cases, we judged that high purity peptides could probably be isolated. By 60 residues, however, the crude peptide product mixtures are probably too heterogeneous to purify to homogeneity by reversed‐phase HPLC. In general, δ‐opioid receptor peptides with a single predicted TM domain were sufficiently soluble to handle postcleavage and to analyze by reversed‐phase HPLC, whereas 1.5 TM domains rendered the peptides too hydrophobic to handle or analyze by standard protocols. Given the challenges of chain assembly, handling, and purification of these peptides, a synthetic strategy that uses ∼ 12 or 13 shorter peptide segments of 20–40 residues each is probably a more feasible approach. © 2006 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 88: 340–349, 2007. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com