Title : N-glycosylation of
prostaglandin endoperoxide synthases-1 and -2 and their orientations in the endoplasmic reticulum
Abstract :
- Using site-directed mutagenesis, we have determined that Asn68, Asn144, and Asn410 of ovine prostaglandin endoperoxide ( PGH) synthase-1 are N-glycosylated
- A fourth consensus N-glycosylation sequence at Asn104 is not glycosylated
- Glycosylation of PGH synthase-1 at Asn410 and at either Asn68 or Asn144 was required for expression of both the cyclooxygenase and the peroxidase activities of the enzyme
- Inactive PGH synthase-1 glycosylation site mutant proteins do not appear to achieve their native conformations
- However, the native enzyme , once in an active conformation, does not appear to require attached carbohydrate for cyclooxygenase or peroxidase activities
- N-Glycosylation consensus sequences corresponding to the three glycosylation sites of ovine PGH synthase-1 are conserved in the deduced amino acid sequences of PGH synthases-2
- Using site-directed mutagenesis, we determined that there is an additional site of N-glycosylation in murine PGH synthase-2 located at Asn580
- This site is N-glycosylated in about 50% of PGH synthase-2 molecules, resulting in two peptide bands on SDS-polyacrylamide gel electrophoresis (72 and 74 kDa)
- Glycosylation of PGH synthase-2 is necessary for expression of enzyme activity, but glycosylation of PGH synthase-2 at Asn580 per se does not affect activity
- Assuming that the N-glycosylation sites of PGH synthase-1 are on the luminal side of the endoplasmic reticulum (ER), and that the site of tryptic cleavage of ovine PGH synthase-1 ( Arg277 ) is on the cytoplasmic side of the ER, we propose that both the NH2 and COOH termini of PGH synthase-1 are located in the lumen of the ER and that there are two transmembrane domains located between Asn144 and Arg277 and between Arg277 and Asn410 , respectively
- A similar orientation is predicted for PGH synthase-2
Output (sent_index, trigger,
protein,
sugar,
site):
- 1. N-glycosylated, , -, -, Asn68, Asn144, and Asn410
- 10. N-glycosylation, , PGH synthase-1, -, sites
- 2. glycosylated, , -, -, sequence
- 3. Glycosylation, , PGH synthase-1, -, Asn144
- 3. Glycosylation, , PGH synthase-1, -, Asn410
- 3. Glycosylation, , PGH synthase-1, -, Asn68
- 4. glycosylation, , -, -, site
- 6. glycosylation, , PGH synthase-1, -, sites
- 7. N-glycosylation, , PGH synthase-2, -, site
- 8. N-glycosylated, , -, -, site
- 9. Glycosylation, , PGH synthase-2, -, -
- 9. glycosylation, , -, -, Asn580
- 9. glycosylation, , PGH synthase-2, -, Asn580
Output(Part-Of) (sent_index,
protein,
site):
- 10. PGH synthase-1, Arg277
- 10. PGH synthase-1, sites
- 10. PGH synthase-1, termini
- 6. PGH synthase-1, sites
- 7. PGH synthase-2, site
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 1. prostaglandin endoperoxide synthases-1, -, Asn68, Asn144, and Asn410
- 3. PGH synthase-1, -, Asn144
- 3. PGH synthase-1, -, Asn410
- 3. PGH synthase-1, -, Asn68
- 9. PGH synthase-2, -, Asn580