Title : Site-specific N-glycosylation analysis of human
factor XI : Identification of a noncanonical NXC
glycosite
Abstract :
- Human factor XI ( hFXI ) is a 160-kDa disulphide-linked homodimer zymogen involved in the coagulation cascade
- Its deficiency results in bleeding diathesis referred to as hemophilia C. hFXI bears five N-glycosylation consensus sites per monomer, N72 , N108 , N335 on the heavy chain and N432 , N473 on the light chain
- This study reports the first in-depth glycosylation analysis of hFXI based on advanced MS approaches
- Hydrophilic interaction LC and MS characterization and quantification of the N-glycans showed that the two major forms are complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures (A2 S2 , 66%)
- Minor triantennary structures (A3 S3 F, ∼1.5%; A3 S3 , ∼2%) were also identified
- MS analyses of intact hFXI revealed full occupation of two of the three heavy-chain glycosites and almost full-site occupancy of the light chain
- Analysis of hFXI glycopeptides by LC-MS/MS enabled site-specific glycan profiling and occupancy
- It was evidenced that N335 was not glycosylated and that N72 and N108 were fully occupied, whereas N432 and N473 were occupied at about 92 and 95%, respectively
- We also identified a new glycosite of the noncanonical format NXC at N145 , occupied at around 5%
- These data provide valuable structural information useful to understand the potential roles of N-glycosylation on hFXI function and could serve as a structural reference
Output (sent_index, trigger,
protein,
sugar,
site):
- 0. N-glycosylation, , factor XI, -, -
- 0. glycosite, , -, -, glycosite
- 2. N-glycosylation, , -, -, N432 , N473
- 2. N-glycosylation, , -, -, N72 , N108 , N335
- 2. N-glycosylation, , -, -, sites
- 3. glycosylation, , hFXI, -, -
- 4. A2, , -, complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures, A2-S2
- 4. bis-α2,6-sialylated, , -, complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures, A2-S2
- 4. mono-α2,6-sialylated, , -, complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures, A2-S2
- 6. glycosites, , -, -, glycosites
- 7. glycopeptides, , -, -, glycopeptides
- 8. glycosylated, , -, -, N335
- 8. occupied, , -, -, N432 and N473
- 8. occupied, , -, -, N72 and N108
- 9. glycosite, , -, -, glycosite
Output(Part-Of) (sent_index,
protein,
site):
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 2. factor XI, -, N432 , N473
- 2. factor XI, -, N72 , N108 , N335
- 4. factor XI, complex biantennary mono-α2,6-sialylated (A2 S1 , 20%) and bis-α2,6-sialylated structures, A2-S2
- 8. factor XI, -, N335
- 8. factor XI, -, N432 and N473
- 8. factor XI, -, N72 and N108