N‐glycosylation proteomic characterization and cross‐species comparison of milk whey proteins from dairy animals

Abstract

Glycosylated proteins in milk have been implicated in multiple biological roles. However, the N-glycoprotein components and their complexity in milk whey from dairy animals are not well characterized. Here, a modified proteomics approach consisting of N-glycopeptide enrichment and identification was used to map the N-glycoproteome profile of milk whey from Holstein and Jersey cows, buffaloes, yaks, goats, camels, and horses. A total of 233 N-glycosylation sites, corresponding to 147 N-glycoproteins, were detected in the studied animals. Most of the identified N-glycosylation sites were not characterized in the database and were considered as unknown. Functional analysis of the identified glycoproteins demonstrated that response to stimulus was the most abundant GO category shared in the studied animals according to their annotation. Lysosome, glycosaminoglycan degradation, and extracellular matrix-receptor interaction pathways were shared between Holstein and Jersey cows, and yaks. N-glycoprotein components of milk whey from Holstein and Jersey cows, buffaloes, yaks, and goats were more similar to each other than to those of camels and horses. These results significantly extend the number of known N-glycosylation sites and reveal in-depth composition and potential functions of N-glycoproteins in milk whey, which in turn provides insights to further explore N-glycoprotein biosynthesis in the studied animals.

Glycoproteomics Analysis Reveals Differential Expression of Site-Specific Glycosylation in Human Milk Whey during Lactation

“…The glycoproteome of milk whey has been analyzed by LC–MS/MS, and most studies have identified glycosites by analyzing enzymatically deglycosylated peptides or identified glycans that are enzymatically released from N-glycoproteins in human and other mammalian milks. ,,,− For example, Cao et al applied lectin to enrich the glycopeptides and used PNGase F to remove the N-glycans from the enriched glycopeptides. They then analyzed the resulting deglycosyated peptides by LC–MS/MS, identifying 63 glycoproteins with 110 N-glycosites and 53 glycoproteins with 91 N-glycosites in colostrum and mature milk whey, respectively.…”

Comparative Analysis of Whey N-Glycoproteins in Human Colostrum and Mature Milk Using Quantitative Glycoproteomics

“…Recently, major whey N-glycoproteins in dairy milk were reported to be involved in response to stimulus. 33 In the current study, the differentially expressed whey N-glycoproteins in human colostrum and mature milk were found to be enriched mainly in localization, extracellular region part, and sulfur compound binding. It is noteworthy that 16 differentially expressed N-glycoproteins with 36 glycosylation sites were related to immune system process, of which 9 glycoproteins contained upregulated sites, 3 contained downregulated sites, and 4 contained upregulated and downregulated sites (Table 1).…”

“…30,31 Most recently, large-scale identification of N-glycoproteomes were performed in MFGM from humans and seven mammals and in whey from dairy milk. 32,33 Although some researches has focused on milk glycoproteins, the largescale identification and quantification of whey N-glycosylation sites in human milk by glycoproteomics has not been well elucidated.…”

Proteomic analysis of differentially expressed whey proteins in Guanzhong goat milk and Holstein cow milk by iTRAQ coupled with liquid chromatography-tandem mass spectrometry

“…These differentially expressed proteins related to glycoproteins may play a role in protecting infants from disease differently between goat milk and cow milk. Yang et al (2017b) studied the N-glycoproteome profile of whey from dairy animals, and found that goat milk whey shared only 32 glycosylated proteins (44.4%) with cow milk whey.…”