Evidence for 4-hydroxyproline in viral proteins. Characterization of a viral prolyl 4-hydroxylase and its peptide substrates

J Biol Chem. 1999 Aug 6;274(32):22131-4. doi: 10.1074/jbc.274.32.22131.


4-Hydroxyproline, the characteristic amino acid of collagens and collagen-like proteins in animals, is also found in certain proline-rich proteins in plants but has been believed to be absent from viral and bacterial proteins. We report here on the cloning and characterization from a eukaryotic algal virus, Paramecium bursaria Chlorella virus-1, of a 242-residue polypeptide, which shows distinct sequence similarity to the C-terminal half of the catalytic alpha subunits of animal prolyl 4-hydroxylases. The recombinant polypeptide, expressed in Escherichia coli, was found to be a soluble monomer and to hydroxylate both (Pro-Pro-Gly)(10) and poly(L-proline), the standard substrates of animal and plant prolyl 4-hydroxylases, respectively. Synthetic peptides such as (Pro-Ala-Pro-Lys)(n), (Ser-Pro-Lys-Pro-Pro)(5), and (Pro-Glu-Pro-Pro-Ala)(5) corresponding to proline-rich repeats coded by the viral genome also served as substrates. (Pro-Ala-Pro-Lys)(10) was a particularly good substrate, with a K(m) of 20 microM. The prolines in both positions in this repeat were hydroxylated, those preceding the alanines being hydroxylated more efficiently. The data strongly suggest that P. bursaria Chlorella virus-1 expresses proteins in which many prolines become hydroxylated to 4-hydroxyproline by a novel viral prolyl 4-hydroxylase.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Cloning, Molecular
  • Escherichia coli / genetics
  • Hydroxylation
  • Hydroxyproline / biosynthesis*
  • Molecular Sequence Data
  • Open Reading Frames
  • Peptides / metabolism
  • Phycodnaviridae / enzymology*
  • Procollagen-Proline Dioxygenase / genetics
  • Procollagen-Proline Dioxygenase / isolation & purification*
  • Procollagen-Proline Dioxygenase / metabolism
  • Proline / metabolism*
  • Recombinant Proteins / isolation & purification
  • Recombinant Proteins / metabolism
  • Repetitive Sequences, Amino Acid
  • Sequence Homology, Amino Acid
  • Viral Proteins / genetics
  • Viral Proteins / isolation & purification*
  • Viral Proteins / metabolism


  • Peptides
  • Recombinant Proteins
  • Viral Proteins
  • Proline
  • Procollagen-Proline Dioxygenase
  • Hydroxyproline