Changes in the proteome of yolk granules induced by freezing: Label-free proteomics and molecular dynamics simulation...

Freezing-induced changes of the egg yolk granules significantly compromises the functional properties of freeze-thawed egg yolk. To elucidate the molecular mechanisms underlying this phenomenon, this study employed an integrative approach combining proteomic analysis and molecular dynamics (MD) simulations to investigate freezing-induced proteome changes in egg yolk granules. The results identify low-density lipoprotein (LDL), high-density lipoprotein (HDL), yolk immunoglobulin (IgY), riboflavin-binding protein (RBP), and phosvitin (PV) as key proteins contributors to freezing-related structural alterations. MD simulations conducted at 27 °C and -20 °C revealed that reduced solubility primarily arises from hydrophobic interactions and hydrogen bonding. Critical residues mediating these interactions include VAL/ILE/LEU/GLN/ALA/SER clusters in LDL (e.g., VAL-89, ILE-82, LEU-60), LEU-200/THR-197 in HDL, VAL-374/433 in IgY, THR/SER/LYS/ASN residues (e.g., THR-56, ASN-18) in RBP, and GLU/PRO/LYS/ILE motifs (e.g., GLU-26, ILE-7) in PV. These findings demonstrate that freeze-thaw-induced gelation in egg yolk is governed by synergistic interactions across multiple protein components rather than isolated molecular events.