N-Glycoproteomic Analysis of Chicken Egg Yolk

Abstract

Posttranslational N-glycosylation of food proteins plays a critical role in their structure and function. However, the N-glycoproteome of chicken egg yolk (CEY) has not been studied yet. Glycopeptides hydrolyzed from CEY proteins were enriched, with deglycosylation occurring using PNGase F, and then were identified using a shotgun glycoproteomics strategy. A total of 217 N-glycosylation sites and 86 glycoproteins were identified in CEY, and these glycoproteins are mainly involved in the binding, biological regulation, catalytic activity, and metabolic processes. Among the identified CEY glycoproteins, 22 were recognized as proteases and protease inhibitors, suggesting that a proteinase/inhibitor regulation system exists in CEY; further, 15 were members of the complement and immune systems, which provide protection against potential threats during hatching.The study provides important structural information about CEY glycoproteins and aids in the understanding of the underlying mechanism of embryo development as well as changes in CEY functional characteristics during storage and processing.

Comparative proteomic analysis of hen egg yolk plasma proteins during embryonic development

“…Five ovotransferrin spots were identified through 2‐DE gel analysis in egg white (Guerin‐Dubiard, ). As shown in Figures and , these five ovotransferrin spots were differentiated distinctly and one of them (spot 18) was identified with a higher MW than theoretical value (Table ), indicating possible glycosylation (Geng, Xie, Wang, Kaustav, Qiu, & Ma, ). Ovotransferrin is a highly abundant protein in egg white.…”

Analysis of tartary buckwheat ( Fagopyrum tataricum ) seed proteome using offline two‐dimensional liquid chromatography and tandem mass spectrometry

“…The extracted TBS total protein samples (200 μg) were reduced with 5 mmol/L dithiothreitol at 56°C for 30 min, and then alkylated with 10 mmol/L of iodoacetamide for 15 min at room temperature in the dark. Subsequently, the TBS total protein was digested using trypsin (Sigma‐Aldrich) at 1:50 trypsin‐to‐protein mass ratio for the first digestion overnight and 1:100 trypsin‐to‐protein mass ratio for a second 4‐hr digestion (Geng et al, ; Wang et al, ). Three biological replicates were performed to ensure protein reproducibility.…”

“…Peptides were then selected for MS/MS using a dynamically excluded time of 30 s, and the fragments were detected at a resolution of 17,500. Automatic gain control was set at 5E4 (Geng, Wang, Liu, Jin, & Ma, 2017;Geng et al, 2018;Wang et al, 2019).…”

In‐depth mapping of the proteome of Tibetan pig tenderloin ( longissimus dorsi ) using offline high‐pH reversed‐phase fractionation and LC‐MS/MS

“…The peptides were loaded onto a Reprosil‐Pur C18 reverse‐phase analytical column (1.9 μm particles, inner diameter 75 µm, 15 cm length) and separated with a gradient of buffer A (2% acetonitrile with 0.1% formic acid) and buffer B (90% acetonitrile with 0.1% formic acid) at a flow rate of 300 nl/min. Buffer B increased from 9% to 25% in 24 min and increased from 25% to 36% in 8 min, then increased to 80% in 4 min, where it was held for 4 min before decreasing back to 9% B (Geng et al, ; Wang, Xiao, Liu, et al, ).…”

“…Subsequently, the Kyoto Encyclopedia of Genes and Genomes (KEGG, http://www.genome.jp/kegg/pathway.html) database was used to annotate the protein pathway. First, the KEGG online service tool KAAS was used to annotated protein's KEGG database description; then, the annotation results were mapped on the KEGG pathway database using the KEGG online service tool KEGG mapper (Geng et al, ; Wang, Xiao, Liu, et al, ).…”