Membrane-anchored prolyl hydroxylase with an export signal from the endoplasmic reticulum

• Published in: The Plant journal : for cell and molecular biology Vol. 41; no. 1; pp. 81 - 94
• Main Authors: Yuasa, K, Toyooka, K, Fukuda, H, Matsuoka, K
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 2005; Blackwell Science; Blackwell Publishing Ltd
• Subjects: Amino Acid Sequence; Amino acids; Arabidopsis - genetics; Biological and medical sciences; complementary DNA; cultured cells; endoplasmic reticulum; Endoplasmic Reticulum - enzymology; Endoplasmic Reticulum - metabolism; Enzymes; export signal; expressed sequence tags; Fundamental and applied biological sciences. Psychology; Gene expression; Golgi apparatus; Golgi Apparatus - enzymology; Golgi Apparatus - metabolism; membrane protein; membrane proteins; Metabolism; Molecular Sequence Data; Nicotiana - genetics; Nicotiana - metabolism; Nicotiana tabacum; Oryza - genetics; oxygenases; Plant physiology and development; Plant populations; plant proteins; procollagen-proline dioxygenase; Procollagen-Proline Dioxygenase - genetics; Procollagen-Proline Dioxygenase - metabolism; prolyl hydroxylase; Proteins; recombinant fusion proteins; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Rice; secretory pathway; Sequence Homology, Amino Acid; Signal Transduction; tissue distribution; Tobacco; tobacco BY‐2 cells; Heterologous system; Enzyme; Immunocytochemistry; prolyl hydroxylase; Homology; secretory pathway; Gene expression; Nicotiana tabacum; export signal; Characterization; tobacco BY-2 cells; Dicotyledones; Angiospermae; membrane protein; Spermatophyta; Oxidoreductases; Solanaceae; Expressed sequence tag; Procollagen-proline dioxygenase
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/j.1365-313X.2004.02279.x

We cloned a novel prolyl 4-hydroxylase (PH; EC 1.14.11.2) homolog cDNA from tobacco (Nicotiana tabacum) BY-2 cells based on expression sequence tag information. Like other PHs, this tobacco PH polypeptide has two conserved histidine residues, and it comprises 286 amino acids with a calculated molecular mass of 32 kDa. Interestingly, this protein and homologs in Arabidopsis and rice have predicted transmembrane sequences in their N-terminal regions. This PH homolog was expressed in BY-2 cells as a His-tagged protein, and the expressed protein showed PH activity. Incubation of membranes with high salt, urea, and protease with or without detergents indicated that this protein is an integral membrane protein with a type II configuration. Its membrane-anchored nature is specific for plants because no integral membrane PH has been found in animals. A membrane fractionation study and immunocytochemical studies indicate that this protein localizes in both the endoplasmic reticulum (ER) and Golgi apparatus. Analysis of this protein fused to green fluorescent protein indicated that basic amino acids in the cytoplasmic, N-terminal region of the PH play a role in its export from the ER.