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

Abstract

Glycosylation is a ubiquitous post-translational protein modification that plays a substantial role in various processes. However, whey glycoproteins in human milk have not been completely profiled. Herein, we used quantitative glycoproteomics to quantify whey N-glycosylation sites and their alteration in human milk during lactation; 110 N-glycosylation sites on 63 proteins and 91 N-glycosylation sites on 53 proteins were quantified in colostrum and mature milk whey, respectively. Among these, 68 glycosylation sites on 38 proteins were differentially expressed in human colostrum and mature milk whey. These differentially expressed N-glycoproteins were highly enriched in "localization", "extracellular region part", and "modified amino acid binding" according to gene ontology annotation and mainly involved in complement and coagulation cascades pathway. These results shed light on the glycosylation sites, composition and biological functions of whey N-glycoproteins in human colostrum and mature milk, and provide substantial insight into the role of protein glycosylation during infant development.

Meta‐omics reveal microbial assortments and key enzymes in bean sauce mash, a traditional fermented soybean product

“…Gene ontology (GO) annotation (http://www.geneontology.org/) was employed to analyze the function of proteins that were identified. The GO categories for which the P value was smaller than 0.05 were applied for analysis http://www.ncbi.nlm.nih.gov/COG/…”

“…The GO categories for which the P value was smaller than 0.05 were applied for analysis. 25 http://www.ncbi.nlm.nih .gov/COG/http://www Total DNA extraction Five sites of each bean sauce mash were collected from different parts of the whole sample, contains the exterior or interior of the bean sauce mash; 2 g was collected from each site, and DNA extraction was performed using the Soil DNA Kit (Tiangen, Beijing, China), according to the manufacturer's instructions. The DNA extractions from five different locations of the sample were combined in a 1.5 mL sterile centrifuge tube and mixed well.…”

Label-Free Mass Spectrometry-Based Quantitative Proteomics Analysis of Serum Proteins During Early Pregnancy in Jennies (Equus asinus)

“…LC-ESI-MS/MS was performed on a Q-Exactive Plus mass spectrometer coupled with an EASY 1200 nano-LC System (Thermo Fisher Scientific, Bremen, Germany). LC-ESI-MS/MS settings were the same as those stated in a previous study (22). Briefly, 3 µg of the peptide mixture was first loaded onto a trap column (100 µm inner diameter, 20 mm long, 5 µm resin, C18, Dr. Maisch GmbH, Ammerbuch, Germany) in buffer A (0.1% formic acid in water).…”

Mapping human N-linked glycoproteins and glycosylation sites using mass spectrometry

“…From CSF, a total of >1,600 glycosites from ~700 glycoproteins were identified in health and disease-related samples using multiple strategies, including gel separation, reversed phase-anion exchange and hydrazidebased glycopeptide capture [54][55][56][57]. In human milk, 240 N-glycosylation sites from 114 glycoproteins were identified in human colostrum and mature milk using HILIC enrichment, deglycosylation via PNGase F treatment and MS analysis [58][59][60]. These data might provide insight into the potential applications of some N-glycoproteins in infant formulae at different stages of development.…”