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Site-specific protein modifications through pyrroline-carboxy-lysine residues

Academic Article
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Overview

authors

  • Ou, W. J.
  • Uno, T.
  • Chiu, H. P.
  • Grunewald, J.
  • Cellitti, S. E.
  • Crossgrove, T.
  • Hao, X. S.
  • Fan, Q.
  • Quinn, L. L.
  • Patterson, P.
  • Okach, L.
  • Jones, D. H.
  • Lesley, Scott
  • Brock, A.
  • Geierstanger, B. H.

publication date

  • 2011

journal

  • Proceedings of the National Academy of Sciences of the United States of America  Journal

abstract

  • Pyrroline-carboxy-lysine (Pcl) is a demethylated form of pyrrolysine that is generated by the pyrrolysine biosynthetic enzymes when the growth media is supplemented with D-ornithine. Pcl is readily incorporated by the unmodified pyrrolysyl-tRNA/tRNA synthetase pair into proteins expressed in Escherichia coli and in mammalian cells. Here, we describe a broadly applicable conjugation chemistry that is specific for Pcl and orthogonal to all other reactive groups on proteins. The reaction of Pcl with 2-amino-benzaldehyde or 2-amino-acetophenone reagents proceeds to near completion at neutral pH with high efficiency. We illustrate the versatility of the chemistry by conjugating Pcl proteins with poly(ethylene glycol)s, peptides, oligosaccharides, oligonucleotides, fluorescence, and biotin labels and other small molecules. Because Pcl is genetically encoded by TAG codons, this conjugation chemistry enables enhancements of the pharmacology and functionality of proteins through site-specific conjugation.

subject areas

  • Culture Media
  • Escherichia coli
  • Lysine
  • Nuclear Magnetic Resonance, Biomolecular
  • Proteins
  • Pyrroles
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Research

keywords

  • amber suppression
  • desmethylpyrrolysine
  • protein engineering
  • protein medicinal chemistry
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Identity

PubMed Central ID

  • PMC3127931

International Standard Serial Number (ISSN)

  • 0027-8424

Digital Object Identifier (DOI)

  • 10.1073/pnas.1105197108

PubMed ID

  • 21670250
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Additional Document Info

start page

  • 10437

end page

  • 10442

volume

  • 108

issue

  • 26

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