Protein-glutaminase-derived small peptides mitigate post-acidification in yogurt by activating the arginine-fatty acid metabolic pathway

Post-acidification remains a persistent quality defect in yogurt production. Enzymatic intervention can modulate it by regulating microbial metabolism. This study evaluated 0.00–0.12 U/mL protein glutaminase (PG) effects on post-acidification through fermentation kinetics and organic acid analysis. Notably, 0.06 U/mL PG controlled acidification, showing a Δ7 °T acidity increase and 9.06 mg/g lactic acid over 14-day storage, enhancing water holding capacity, viscosity, texture, and casein matrix compactness. Metabolomic and peptidomic analyses showed PG at this dose increased peptide concentration to 24.31 μg/μL, modifying 26 amino acids/small peptides (54.54% tripeptides). After 14 days, enriched amino acids were linked to amino acid and fatty acid biosynthesis pathways. Gene-level investigations revealed upregulated arginine ( argJ – H , 1.1–17.5-fold) and fatty acid synthesis genes ( accA – fabK , 1.0–6.0-fold) in starter cultures, increasing membrane UFA/SFA ratios. Collectively, 0.06 U/mL PG treatment enhanced polypeptide levels, activated pathways for arginine, polyamines, and fatty acids, improved membrane fluidity, reduced post-acidification, and enhanced yogurt quality. • 0.06 U/mL PG improved post-acidification by optimizing fermentation kinetics. • 0.06 U/mL PG enhanced yogurt texture/rheology and compacted casein matrices. • 0.06 U/mL PG altered milk matrix metabolites and increased peptide levels. • Amino acids and small peptides were enriched in amino and fatty acid pathways. • Arginine-fatty acid gene upregulation raised UFA/SFA ratio, reducing post-acidification.