N‑Glycoproteomics of Human Seminal Plasma Glycoproteins

Seminal
plasma aids sperm by inhibiting premature capacitation,
helping in the intracervical transport and formation of an oviductal
sperm reservoir, all of which appear to be important in the fertilization
process. Epitopes such as Lewis x and y are known to be present on
seminal plasma glycoproteins, which can modulate the maternal immune
response. It is suggested by multiple studies that seminal plasma
glycoproteins play, largely undiscovered, important roles in the process
of fertilization. We have devised a strategy to analyze glycopeptides
from a complex, unknown mixture of protease-digested proteins. This
analysis provides identification of the glycoproteins, glycosylation
sites, glycan compositions, and proposed structures from the original
sample. This strategy has been applied to human seminal plasma total
glycoproteins. We have elucidated glycan compositions and proposed
structures for 243 glycopeptides belonging to 73 N-glycosylation sites
on 50 glycoproteins. The majority of the proposed glycan structures
were complex type (83%) followed by high-mannose (10%) and then hybrid
(7%). Most of the glycoproteins were either sialylated, fucosylated,
or both. Many Lewis x/a and y/b epitopes bearing glycans were found,
suggesting immune-modulating epitopes on multiple seminal plasma glycoproteins.
The study also shows that large scale N-glycosylation mapping is achievable
with current techniques and the depth of the analysis is roughly proportional
to the prefractionation and complexity of the sample.