Identification of Lactoferricin B Intracellular Targets Using an Escherichia Coli Proteome Chip

Overview

Journal PLoS One

Specialties General Medicine
Science

Date 2011 Dec 14

PMID 22164243

Citations 21

Authors

Yu-Hsuan Tu

Yu-Hsuan Ho

Ying-Chih Chuang

Po-Chung Chen

Chien-Sheng Chen

Affiliations

Soon will be listed here.

Abstract

Lactoferricin B (LfcinB) is a well-known antimicrobial peptide. Several studies have indicated that it can inhibit bacteria by affecting intracellular activities, but the intracellular targets of this antimicrobial peptide have not been identified. Therefore, we used E. coli proteome chips to identify the intracellular target proteins of LfcinB in a high-throughput manner. We probed LfcinB with E. coli proteome chips and further conducted normalization and Gene Ontology (GO) analyses. The results of the GO analyses showed that the identified proteins were associated with metabolic processes. Moreover, we validated the interactions between LfcinB and chip assay-identified proteins with fluorescence polarization (FP) assays. Sixteen proteins were identified, and an E. coli interaction database (EcID) analysis revealed that the majority of the proteins that interact with these 16 proteins affected the tricarboxylic acid (TCA) cycle. Knockout assays were conducted to further validate the FP assay results. These results showed that phosphoenolpyruvate carboxylase was a target of LfcinB, indicating that one of its mechanisms of action may be associated with pyruvate metabolism. Thus, we used pyruvate assays to conduct an in vivo validation of the relationship between LfcinB and pyruvate level in E. coli. These results showed that E. coli exposed to LfcinB had abnormal pyruvate amounts, indicating that LfcinB caused an accumulation of pyruvate. In conclusion, this study successfully revealed the intracellular targets of LfcinB using an E. coli proteome chip approach.

Citing Articles

Testing Antimicrobial Properties of Human Lactoferrin-Derived Fragments.

Ostrowka M, Duda-Madej A, Pietluch F, Mackiewicz P, Gagat P Int J Mol Sci. 2023; 24(13).

PMID: 37445717 PMC: 10342102. DOI: 10.3390/ijms241310529.

Design and Evaluation of Short Bovine Lactoferrin-Derived Antimicrobial Peptides against Multidrug-Resistant .

Mishra B, Felix L, Basu A, Kollala S, Chhonker Y, Ganesan N Antibiotics (Basel). 2022; 11(8).

PMID: 36009954 PMC: 9404989. DOI: 10.3390/antibiotics11081085.

The Role of Antimicrobial Peptides as Antimicrobial and Antibiofilm Agents in Tackling the Silent Pandemic of Antimicrobial Resistance.

Lopes B, Hanafiah A, Nachimuthu R, Muthupandian S, Md Nesran Z, Patil S Molecules. 2022; 27(9).

PMID: 35566343 PMC: 9105241. DOI: 10.3390/molecules27092995.

Short peptides conjugated to non-peptidic motifs exhibit antibacterial activity.

Ardila-Chantre N, Hernandez-Cardona A, Pineda-Castaneda H, Estupinan-Torres S, Lucia Leal-Castro A, Fierro-Medina R RSC Adv. 2022; 10(49):29580-29586.

PMID: 35521126 PMC: 9055962. DOI: 10.1039/d0ra05937d.

Multiple roles of ribosomal antimicrobial peptides in tackling global antimicrobial resistance.

Luong H, Ngan H, Thi Phuong H, Quoc T, Tung T R Soc Open Sci. 2022; 9(1):211583.

PMID: 35116161 PMC: 8790363. DOI: 10.1098/rsos.211583.

References

Thao S, Chen C, Zhu H, Escalante-Semerena J . Nε-lysine acetylation of a bacterial transcription factor inhibits Its DNA-binding activity. PLoS One. 2011; 5(12):e15123. PMC: 3013089. DOI: 10.1371/journal.pone.0015123. View

Chen C, Sullivan S, Anderson T, Tan A, Alex P, Brant S . Identification of novel serological biomarkers for inflammatory bowel disease using Escherichia coli proteome chip. Mol Cell Proteomics. 2009; 8(8):1765-76. PMC: 2722769. DOI: 10.1074/mcp.M800593-MCP200. View

Saka K, Tadenuma M, Nakade S, Tanaka N, Sugawara H, Nishikawa K . A complete set of Escherichia coli open reading frames in mobile plasmids facilitating genetic studies. DNA Res. 2005; 12(1):63-8. DOI: 10.1093/dnares/12.1.63. View

Chen C, Korobkova E, Chen H, Zhu J, Jian X, Tao S . A proteome chip approach reveals new DNA damage recognition activities in Escherichia coli. Nat Methods. 2007; 5(1):69-74. PMC: 2576280. DOI: 10.1038/nmeth1148. View

Boal A, Cotruvo Jr J, Stubbe J, Rosenzweig A . Structural basis for activation of class Ib ribonucleotide reductase. Science. 2010; 329(5998):1526-30. PMC: 3020666. DOI: 10.1126/science.1190187. View

Cudic M, Otvos Jr L . Intracellular targets of antibacterial peptides. Curr Drug Targets. 2002; 3(2):101-6. DOI: 10.2174/1389450024605445. View

Zhu X, Gerstein M, Snyder M . ProCAT: a data analysis approach for protein microarrays. Genome Biol. 2006; 7(11):R110. PMC: 1794587. DOI: 10.1186/gb-2006-7-11-r110. View

Jordan A, Pontis E, Atta M, Krook M, Gibert I, Barbe J . A second class I ribonucleotide reductase in Enterobacteriaceae: characterization of the Salmonella typhimurium enzyme. Proc Natl Acad Sci U S A. 1994; 91(26):12892-6. PMC: 45546. DOI: 10.1073/pnas.91.26.12892. View

Knight S, Umezawa N, Lee H, Gellman S, Kay B . A fluorescence polarization assay for the identification of inhibitors of the p53-DM2 protein-protein interaction. Anal Biochem. 2002; 300(2):230-6. DOI: 10.1006/abio.2001.5468. View

10.

Jameson D, Seifried S . Quantification of protein-protein interactions using fluorescence polarization. Methods. 1999; 19(2):222-33. DOI: 10.1006/meth.1999.0853. View

11.

Maere S, Heymans K, Kuiper M . BiNGO: a Cytoscape plugin to assess overrepresentation of gene ontology categories in biological networks. Bioinformatics. 2005; 21(16):3448-9. DOI: 10.1093/bioinformatics/bti551. View

12.

Brogden K . Antimicrobial peptides: pore formers or metabolic inhibitors in bacteria?. Nat Rev Microbiol. 2005; 3(3):238-50. DOI: 10.1038/nrmicro1098. View

13.

Lueking A, Possling A, Huber O, Beveridge A, Horn M, Eickhoff H . A nonredundant human protein chip for antibody screening and serum profiling. Mol Cell Proteomics. 2003; 2(12):1342-9. DOI: 10.1074/mcp.T300001-MCP200. View

14.

Allen M, Reeves J, Mellor G . High throughput fluorescence polarization: a homogeneous alternative to radioligand binding for cell surface receptors. J Biomol Screen. 2000; 5(2):63-9. DOI: 10.1177/108705710000500202. View

15.

Chandra H, Reddy P, Srivastava S . Protein microarrays and novel detection platforms. Expert Rev Proteomics. 2011; 8(1):61-79. DOI: 10.1586/epr.10.99. View

16.

Cline M, Smoot M, Cerami E, Kuchinsky A, Landys N, Workman C . Integration of biological networks and gene expression data using Cytoscape. Nat Protoc. 2007; 2(10):2366-82. PMC: 3685583. DOI: 10.1038/nprot.2007.324. View

17.

Haukland H, Ulvatne H, Sandvik K, Vorland L . The antimicrobial peptides lactoferricin B and magainin 2 cross over the bacterial cytoplasmic membrane and reside in the cytoplasm. FEBS Lett. 2001; 508(3):389-93. DOI: 10.1016/s0014-5793(01)03100-3. View

18.

Lundblad J, Laurance M, Goodman R . Fluorescence polarization analysis of protein-DNA and protein-protein interactions. Mol Endocrinol. 1996; 10(6):607-12. DOI: 10.1210/mend.10.6.8776720. View

19.

Zhu J, Shimizu K . Effect of a single-gene knockout on the metabolic regulation in Escherichia coli for D-lactate production under microaerobic condition. Metab Eng. 2005; 7(2):104-15. DOI: 10.1016/j.ymben.2004.10.004. View

20.

Podda E, Benincasa M, Pacor S, Micali F, Mattiuzzo M, Gennaro R . Dual mode of action of Bac7, a proline-rich antibacterial peptide. Biochim Biophys Acta. 2006; 1760(11):1732-40. DOI: 10.1016/j.bbagen.2006.09.006. View