Prolyl 3-hydroxylase: partial characterization of the enzyme from rat kidney cortex

Eur J Biochem. 1977 Mar 1;73(2):485-92. doi: 10.1111/j.1432-1033.1977.tb11341.x.


The formation of 3-hydroxyproline was studied with crude rat kidney cortex extract as a source of enzyme and chick embryo tendon protocollagen and procollagen or cartilage protocollagen as a substrate. Synthesis of 3-hydroxyproline was observed with all these substrates and the formation of 3-hydroxyproline ranged up to seven residues per pro-alpha-chain. The highest rate of 3-hydroxylation took place at 20 degrees C and the reaction required Fe2+, O2,2-oxoglutarate and ascorbate. The formation of 3-hydroxyproline was affected by chain length and the conformation of the substrate, in that longer polypeptide chains proved better substrates, while the native triple-helical conformation of protocollagen or procollagen completely prevented the reaction. Formation of 3-hydroxyproline with tendon procollagen as a substrate was not inhibited by antiserum to prolyl 4-hydroxylase or by poly(L-proline) when these substances were used in concentrations which clearly inhibited 4-hydroxyproline formation with tendon protocollagen as a substrate. Furthermore, pure prolyl 4-hydroxylase did not synthesize any 3-hydroxyproline under conditions in which the crude rat kidney cortex enzyme would readily do so. The data thus strongly suggest that prolyl 3-hydroxylase and prolyl 4-hydroxylase are separate enzymes.

MeSH terms

  • Animals
  • Chick Embryo
  • Kidney Cortex / enzymology*
  • Kinetics
  • Peptide Fragments / analysis
  • Procollagen
  • Procollagen-Proline Dioxygenase / metabolism*
  • Protein Conformation
  • Rats
  • Structure-Activity Relationship
  • Temperature
  • Tendons


  • Peptide Fragments
  • Procollagen
  • Procollagen-Proline Dioxygenase