Influencing Mechanisms and Additive-Mediated Modulation of Aggregation Behavior in Papain-Catalyzed Soy Protein Hydrolysis
- 1State Key Laboratory of Food Science and Resources, Jiangnan University, Wuxi 214122, China
- 2Analysis and Testing Center, Jiangnan University, Wuxi 214122, China
- 3Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China
Abstract
Protein aggregation during enzymatic hydrolysis detrimentally impacts product quality. However, due to the inherent complexity of soy protein systems, their precise aggregation mechanisms remain elusive. In this study, papain was employed to hydrolyze soy protein isolate (SPI), as well as 7S and 11S globulins, to elucidate the underlying aggregation mechanisms and explore potential regulatory strategies. The solubility characteristics of the resulting aggregates were evaluated via turbidity measurements (OD660) following treatments with varying concentrations of urea, guanidine hydrochloride, sodium dodecyl sulfate (SDS), and β-mercaptoethanol (β-Me). Amino acid composition and sulfhydryl determination revealed that the turbidity of aggregates decreased with increasing concentrations of urea, guanidine hydrochloride, and SDS, indicating that hydrophobic interactions are a major contributing factor to aggregation. Hydrophobic amino acids such as phenylalanine and leucine were enriched in aggregates compared with the supernatant, further supporting the association of hydrophobic interactions. Sulfhydryl analysis showed that total sulfhydryl content increased in hydrolyzed 11S but decreased in 7S, with higher sulfhydryl levels detected in precipitates than in supernatants, suggesting that disulfide bonds exhibit an auxiliary contribution to 11S aggregation. To regulate aggregation, Na2SO3 (0.1%), cysteine, or SDS was added prior to enzyme inactivation. Among these, SDS produced the most significant improvement in supernatant yield (18.77%, 12.57%, and 10.73% for SPI, 7S, and 11S, respectively), primarily by disrupting hydrophobic interactions. Cysteine, as a strong reducing agent, was more effective in suppressing 11S aggregation through the targeted cleavage of disulfide bonds. Tricine-SDS-PAGE confirmed that aggregates were mainly composed of small peptides linked through hydrophobic interactions, with disulfide bonds providing partial stabilization, while both SDS and cysteine effectively inhibited aggregate formation. Overall, hydrophobic interactions were identified as a major correlative factor driving soy protein aggregation during enzymatic hydrolysis, with disulfide bonds playing an auxiliary role in 11S aggregation. These findings provide a robust theoretical basis for the regulation of protein hydrolysate aggregation.
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BibTeX
@article{pan2026influencing,
title = {{Influencing Mechanisms and Additive-Mediated Modulation of Aggregation Behavior in Papain-Catalyzed Soy Protein Hydrolysis}},
author = {Hongyang Pan and Zhaojun Wang and Xiaofang Chu and Jie Chen},
journal = {Molecules},
year = {2026},
month = oct,
doi = {10.3390/molecules31213864},
url = {https://www.mdpi.com/1420-3049/31/21/3864},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Influencing Mechanisms and Additive-Mediated Modulation of Aggregation Behavior in Papain-Catalyzed Soy Protein Hydrolysis
AU - Hongyang Pan
AU - Zhaojun Wang
AU - Xiaofang Chu
AU - Jie Chen
PY - 2026
DA - 2026/10/26
JO - Molecules
DO - 10.3390/molecules31213864
UR - https://www.mdpi.com/1420-3049/31/21/3864
AB - Protein aggregation during enzymatic hydrolysis detrimentally impacts product quality. However, due to the inherent complexity of soy protein systems, their precise aggregation mechanisms remain elusive. In this study, papain was employed to hydrolyze soy protein isolate (SPI), as well as 7S and 11S globulins, to elucidate the underlying aggregation mechanisms and explore potential regulatory strategies. The solubility characteristics of the resulting aggregates were evaluated via turbidity measurements (OD660) following treatments with varying concentrations of urea, guanidine hydrochloride, sodium dodecyl sulfate (SDS), and β-mercaptoethanol (β-Me). Amino acid composition and sulfhydryl determination revealed that the turbidity of aggregates decreased with increasing concentrations of urea, guanidine hydrochloride, and SDS, indicating that hydrophobic interactions are a major contributing factor to aggregation. Hydrophobic amino acids such as phenylalanine and leucine were enriched in aggregates compared with the supernatant, further supporting the association of hydrophobic interactions. Sulfhydryl analysis showed that total sulfhydryl content increased in hydrolyzed 11S but decreased in 7S, with higher sulfhydryl levels detected in precipitates than in supernatants, suggesting that disulfide bonds exhibit an auxiliary contribution to 11S aggregation. To regulate aggregation, Na2SO3 (0.1%), cysteine, or SDS was added prior to enzyme inactivation. Among these, SDS produced the most significant improvement in supernatant yield (18.77%, 12.57%, and 10.73% for SPI, 7S, and 11S, respectively), primarily by disrupting hydrophobic interactions. Cysteine, as a strong reducing agent, was more effective in suppressing 11S aggregation through the targeted cleavage of disulfide bonds. Tricine-SDS-PAGE confirmed that aggregates were mainly composed of small peptides linked through hydrophobic interactions, with disulfide bonds providing partial stabilization, while both SDS and cysteine effectively inhibited aggregate formation. Overall, hydrophobic interactions were identified as a major correlative factor driving soy protein aggregation during enzymatic hydrolysis, with disulfide bonds playing an auxiliary role in 11S aggregation. These findings provide a robust theoretical basis for the regulation of protein hydrolysate aggregation.
SN - 1420-3049
ER -