iTRAQ-Based Quantitative Proteomic Analysis of Antibacterial Mechanism of Milk-Derived Peptide BCp12 against Escherichia coli
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials and Chemicals
2.2. Growth Curves of E. coli
2.3. Confocal Laser Scanning Microscopy (CLSM) Assay
2.4. Determination of the Morphologies of E. coli
2.5. Transmission Electron Microscopy (TEM) Assay
2.6. Proteome Analysis of E. coli
2.6.1. Total Protein Extraction
2.6.2. Protein Digestion and iTRAQ Labeling
2.6.3. High pH-RPLC Separation
2.6.4. LC-MS/MS Analysis
2.6.5. Protein Identification
2.7. Molecular Docking Analysis
2.8. Gel Mobility Shift Assay
2.9. Statistical Analysis
3. Results
3.1. The Effect of BCp12 on E. coli Growth
3.2. Localization Analysis of BCp12 in E. coli Cells
3.3. Effect of BCp12 on E. coli Morphologies
3.4. The Effect of BCp12 on the Internal Structure of E. coli Cells (TEM Observation)
3.5. Proteome Analysis
3.5.1. Protein Extraction and Basic Identification Information
3.5.2. LC-MS/MS Analysis
3.5.3. Functional Annotation of Differentially Expressed Proteins
3.5.4. Expression Changes of Cell Membrane-Related Proteins
3.6. Molecular Docking Analysis
3.7. In Vitro Interaction between BCp12 and DNA
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Target Protein | Docking Score (kcal/mol) | Hydrogen Bond Binding Site | Hydrophobic Interaction Binding Site |
---|---|---|---|
3-ketoacyl-CoA thiolase (fadA) | −6.3 | ala-174,pro-133 | tyr-178, phe-394, val-67, gln-69, pro-133, pro-41, val-136 and val-171 |
acyl-CoA dehydrogenase (fadE) | −6.9 | arg-483, asn-429, gly-428, ser-191, thr-185, Glu-270, pro-220 and arg-416 | ille-423, pro-419, leu-261 and phe-216 |
S-(hydroxymethyl)glutathione dehydrogenase (frmA) | −7 | tyr-88, ile-287 and gln-294 | phe-45, val-291, val-289, ile-296, trp-309, phe-314, leu-105 and ile-287 |
Acyltransferase (aas) | −7.5 | phe-344, arg-10, ser-78 and phe-341 | met-85, pro-86, VAL-89, Leu-93, leu-97, phe-146, leu-143, pro-340 and phe-341 |
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Yang, K.; Shi, Y.; Li, Y.; Wei, G.; Zhao, Q.; Huang, A. iTRAQ-Based Quantitative Proteomic Analysis of Antibacterial Mechanism of Milk-Derived Peptide BCp12 against Escherichia coli. Foods 2022, 11, 672. https://doi.org/10.3390/foods11050672
Yang K, Shi Y, Li Y, Wei G, Zhao Q, Huang A. iTRAQ-Based Quantitative Proteomic Analysis of Antibacterial Mechanism of Milk-Derived Peptide BCp12 against Escherichia coli. Foods. 2022; 11(5):672. https://doi.org/10.3390/foods11050672
Chicago/Turabian StyleYang, Kun, Yanan Shi, Yufang Li, Guangqiang Wei, Qiong Zhao, and Aixiang Huang. 2022. "iTRAQ-Based Quantitative Proteomic Analysis of Antibacterial Mechanism of Milk-Derived Peptide BCp12 against Escherichia coli" Foods 11, no. 5: 672. https://doi.org/10.3390/foods11050672