Turnover of prolyl hydroxylase tetramers and the monomer-size protein in chick-embryo cartilaginous bone and lung in vivo

Abstract

The turnover of prolyl hydroxylase and an immunoreactive protein that corresponds in size to the smaller subunit of the enzyme was studied in vivo after-injection of [3H]leucine into 11-day chick embryos. The specific radioactivity and total radioactivity of the monomer-size protein were much higher than those of the enzyme tetramers in the cartilaginous bone at 3 h and 12h after the radioisotope injection, indicating that the monomer-size protein represents precursors rather than degradation products of the enzyme tetramers. Between 24 and 144h after the injection the specific radioactivity and total radioactivity of the two forms of the enzyme protein showed essentially identical decay rates, the observed specific radioactivity of the monomer-size protein being about 120-130% and total radioactivity about 80% of that of the enzyme tetramers. The true half-life, when corrected for dilution caused by tissue growth and re-utilization of the [3H]leucine, was 37.9h for the monomer-size protein and 39.0h for the tetramers. The results obtained in the lung were less reliable owing to high blank radioactivity values in the immunoprecipitation, but even so some definite differences were found between this tissue and the cartilaginous bone. The specific radioactivity of both forms of the enzyme protein at 24h was only about 20-25 % of that in the cartilaginous bone. The total radioactivity of the monomer-size protein in the lung remained about 5 times that of the enzyme tetramers, whereas it was only about 0.8 times that of the tetramers in the cartilaginous bone. As in the cartilaginous bone, the decay rates of both forms of the enzyme protein were essentially identical in the lung, with a true half-life of about 46h. The results suggest that the rate of prolyl hydroxylase synthesis is slower in the lung than in the cartilaginous bone, whereas the degradation rates are fairly similar in these two tissues. The data further suggest that, in the lung at least, a large part of the monomer-size protein became degraded without being converted into enzyme tetramers.

Regulation of collagen post-translational modification in transformed human and chick-embryo cells

“…Interestingly, however, there was a distinct decrease in the fl-subunit-size protein. The association of prolyl hydroxylase subunits to active tetramers probably only occurs after the release of the subunits from the ribosomes, and the fl-subunit-size protein, either the free subunit or its precursor, is utilized in this process (Majamaa et al, 1979;Berg et al, 1980;Kao & Chou, 1980;Kivirikko & Myllyla, 1980). Many observations suggest that in situations with a rapid increase in the rate of collagen synthesis, such as acute experimental liver injury (Risteli et al, , 1978), there is a rapid increase in the synthesis of the a-subunits, this resulting in an elevated concentration of the tetramers and hence elevated enzyme activity (see Kivirikko & Myllyla, 1980).…”

“…Immunoreactive prolyl hydroxylase protein in the human sarcoma cells Immunoreactive prolyl hydroxylase protein is found in isolated cells and in intact tissues in two forms: the active enzyme tetramers (a21J) and an inactive form which corresponds to the smaller enzyme subunit (fl-subunit) both in size and by amino acid and peptide-'map' analysis (see Chen-Kiang et al, 1977;Majamaa et al, 1979;Berg et al, 1980;Kao & Chou, 1980;Kivirikko & Myllyla, 1980). The latter is present in most cells in a large excess over the active tetramers, e.g.…”

Intracellular Enzymes of Collagen Biosynthesis in 3T6 Fibroblasts and Chick‐Embryo Tendon and Cartilage Cells

“…Attempts were also made to determine whether the intracellular enzymes of collagen biosynthesis have relatively short or long half-lives. Recent reports have indicated that prolyl hydroxylase has a relatively long half-life , about 40 h in chick-embryo cartilaginous bone [28], whereas no data are available on the turnover of the other enzymes. Tissue culture media and sera were obtained from Flow Laboratories (Irvine, Scotland), streptomycin sulphate and penicillin G were purchased from Hoechst AG (Frankfurt am Main, F.R.G.…”

Increased mRNAs for procollagens and key regulating enzymes in rat skeletal muscle following downhill running

“…The α-subunit appears to contain the major portion of the catalytic site [20,21]. The β-subunit is produced in excess over the α-subunit [3,17,28] and enters a pool of free subunit before being partially incorporated into the prolyl 4-hydroxylase tetramer [20]. The β-subunit has protein disulphide isomerase activity, both when present as the monomer and when present in the prolyl 4-hydroxylase tetramer.…”