Cloning and Characterization of a Low Molecular Weight Prolyl 4-Hydroxylase from Arabidopsis thaliana
By R. Hieta, J. Myllyharju. Journal of Biological Chemistry, 2002
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4-Hydroxyproline is found in collagens and collagen-like proteins in animals and in many glycoproteins in plants. Animal prolyl 4-hydroxylases (P4Hs) have been cloned and characterized from many sources, but no plant P4H has been cloned so far. We report here that the genome of Arabidopsis thaliana encodes six P4H-like polypeptides, one of which, a 283-residue soluble monomer, was cloned and characterized here as a recombinant protein. Catalytically critical residues identified in animal P4Hs are conserved in this P4H, and their mutagenesis led to complete or almost complete inactivation. The recombinant P4H effectively hydroxylated poly(l-proline) and many synthetic peptides corresponding to proline-rich repeats present in plant glycoproteins and other proteins. Surprisingly, collagen-like peptides were also good substrates, the V max with (Pro-Pro-Gly)10 being similar to that with poly(l-proline). The enzyme acted in this peptide preferentially on prolines in Y positions in theX-Y-Gly triplets. Correspondingly, (Gly-Pro-4Hyp)5 and (Pro-Ala-Gly)5 were poor substrates, with V max values less than 5 and 20% of that obtained with (Pro-Pro-Gly)10, respectively, the K m for the latter also being high. Peptides representing the N- and C-terminal hydroxylation sites present in hypoxia-inducible transcription factor α also served as substrates. As these peptides contain only one proline residue, a poly(l-proline) type II conformation was clearly not required for hydroxylation.
Subfields: Cancer Research, Materials Chemistry, Molecular Biology
View Paper: https://www.semanticscholar.org/paper/d097289b6e8aff514c0101610e398ff8661e5cb4
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Citations in the corpus
- A Bioinformatics Approach to the Identification, Classification, and Analysis of Hydroxyproline-Rich Glycoproteins[W][OA] (2010, background): “Although only four plant P4Hs are cloned and characterized to date (two [P4H1 and P4H2] from Arabidopsis [Hieta and Myllyharju, 2002; Tiainen et al., 2005], one from tobacco [Yuasa et al., 2005], and one from Chlamydomonas [Keskiaho et al., 2007]), 13 P4H genes are predicted to exist for…”
- Fe(II)/α-Ketoglutarate-Dependent Hydroxylases and Related Enzymes (2004, background): “In most cases, the enzyme is comprised of two distinct subunits, the hydroxylase α subunit and a protein disulfide isomerase β subunit which maintains solubility; however, algal and plant prolyl 4-hydroxylases are monomeric proteins (Hieta & Myllyharju, 2002; Kaska et al., 1988).”