Abstract
We cloned a novel prolyl 4-hydroxylase (PH; EC 1.14.11.2) homologue cDNA from tobacco BY-2 cells based on expression sequence tag information. Like other PHs, this tobacco PH polypeptide has two conserved histidine residues. It consisted of 286 amino acids with a calculated molecular mass of 32 kDa. This protein and homologues in Arabidopsis and rice have possible transmembrane regions at their N-terminus. This PH homologue 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 detergents indicated that this protein was an integral membrane protein with a type II configuration. The tobacco PH formed several large protein complexes devoid of protein disulphide isomerase in the membrane.
