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An amalgamation of solid phase peptide synthesis and ribosomal peptide synthesis
Author(s) -
Ottesen Jennifer J.,
BarDagan Maya,
Giovani Baldissera,
Muir Tom W.
Publication year - 2008
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.20810
Subject(s) - semisynthesis , chemical ligation , chemistry , peptide , solid phase synthesis , peptide synthesis , native chemical ligation , combinatorial chemistry , ligation , template , biochemistry , computational biology , stereochemistry , cysteine , nanotechnology , microbiology and biotechnology , enzyme , biology , materials science
Expressed protein ligation (EPL) is a protein semisynthesis technique that allows the site‐specific introduction of unnatural amino acids and biophysical probes into proteins. In the present study, we illustrate the utility of the approach through the generation of two semisynthetic proteins bearing spectroscopic probes. Dihydrofolate reductase containing a single 13 C probe in an active site loop was generated through the ligation of a synthetic peptide‐α‐thioester to a recombinantly generated fragment containing an N‐terminal Cys. Similarly, c‐Crk‐II was assembled by the sequential ligation of three recombinant polypeptide building blocks, allowing the incorporation of 15 N isotopes in the central domain of the protein. These examples showcase the scope of the protein ligation strategy for selective introduction of isotopic labels into proteins, and the protocols described will be of value to those interested in using EPL on other systems. © 2007 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 90:406–414, 2008. This article was originally published online as an acceptedpreprint. The “Published Online” date corresponds to the preprintversion. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com