PMID: 8407981

 

    Legend: Sugar

Title : Core glycosylation of cytochrome P-450(arom)

Abstract :
  1. Evidence for localization of N terminus of microsomal cytochrome P-450 in the lumen
  2. It was found that cytochrome P-450(arom) purified from human placenta microsomes is glycosylated, and the sugar chain was cleaved with endoglycosidase H (Endo H)
  3. The core glycosylation of P-450(arom) was examined with two heterologous expression systems, cultured insect cells and in vitro translation system
  4. The P-450(arom) protein expressed in the insect cells was glycosylated, and the sugar chain was sensitive to Endo H
  5. It was also glycosylated when translated with the wheat germ cell-free system in the presence of rough microsomal membrane, and the sugar chain could be removed by Endo H treatment
  6. Since the P-450(arom) molecule has two potential glycosylation sites ( Asn-12 and Asn-180 ), we replaced each of the 2 asparagine residues with alanine by site-directed mutagenesis and examined the glycosylation of the two mutant proteins in the cell-free system
  7. The core glycosylation did not occur when the Asn-12 residue was mutated, whereas the mutant protein with modified Asn-180 residue was glycosylated
  8. These results demonstrated that the potential glycosylation site ( Asn-12 ) in the N-terminal portion of P-450(arom) is the site of glycosylation
  9. We conclude that the N terminus of P-450(arom) is translocated across the endoplasmic reticulum membrane to be glycosylated at the luminal side