Ref ID : 5930
J.R.A. Leushner and J. Pasternak; Partial purification and characterization of prolyl hydroxylase from the free-living nematode Panagrellus silusiae. Can.J.Zool. 56:159-165, 1978
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Prolyl hydroxylase was isolated from the free-living nematode Panagrellus silusiae by ammonium sulfate precipitation and calcium phosphate gel ion exchange from Triton X-100 treated homogenates. The enzyme preparation was highly purified but not to homogeneity as judged by agarose gel filtration and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Gel filtration indicated that the molecular weight of the enzyme approximated 285 000. Analysis by SDS-acrylamide electrophoresis revealed subunits of ~67 000. Complete enzymic activity required alpha-ketoglutarate, Fe2+, ascorbate, catalase, atmospheric oxygen, and dithiothreitol. This activity was dramatically inhibited by alpha, alpha-dipyridyl, phenanthroline, and polyproline II. The purified enzyme preparation synthesized 50 to 100 µg hydroxyproline per milligram chick protocollagen per hour at 30 degrees C with an estimated Km for the substrate of 80 µg. Thus, a purified prolyl hydroxylase from a simple invertebrate possesses many of the properties of the prolyl hydroxylases of various vertebrates.