Characterization of activatable form of prolyl hydroxylase in L929 fibroblasts.
Prolyl hydroxylase (prolyl-glycyl-peptide, 2-oxoglutarate : oxygen oxidoreductase, EC 1.14.11.2) activity in a sonicated preparation of early log-phase L929 cells could be increased 3-4-times by preincubation of the sonicate with all cofactors of proline hydroxylation, such as ascorbate, Fe2+ and alpha-ketoglutarate. An "activatable" form of the enzyme is produced in these cells due to a deficiency of one of the cofactors in these cultures. The activatable form is found to be different from the active enzyme with respect to its stability to heat and dithiothreitol denaturation. The activatable form has different ionic properties and could be separated from the active enzyme by DEAE-Sephadex chromatography. The available evidence suggests that the activatable form is a tight complex produced by the enzyme with underhydroxylated collagen, the latter being produced by a cofactor deficiency in the cells. Activation of this complex follows the hydroxylation of the substrate and its subsequent release from the bound enzyme.
D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D002478:Cells, Cultured; D004229:Dithiothreitol / Q000494:pharmacology; D004355:Drug Stability; D004789:Enzyme Activation / Q000187:drug effects; D005347:Fibroblasts / Q000201:enzymology*; D006358:Hot Temperature; D007501:Iron / Q000494:pharmacology; D007656:Ketoglutaric Acids / Q000378:metabolism; D008214:Lymphocytes / Q000201:enzymology; D051379:Mice; D009131:Muscle, Smooth, Vascular / Q000201:enzymology; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism*; D011392:Proline / Q000031:analogs & derivatives; D011817:Rabbits
D007656:Ketoglutaric Acids; D011392:Proline; D007501:Iron; D011393:Procollagen-Proline Dioxygenase; D001205:Ascorbic Acid; D004229:Dithiothreitol