Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Antibacterial activity and mechanism of optimized THPA conjugated dipeptides against methicillin-resistant Staphylococcus aureus

The Journal of Antibiotics volume 79, pages 15–29 (2026)Cite this article

Subjects

Abstract

Despite decades of investigation, multidrug-resistant bacterial resistance has become the leading health concern globally related to MRSA infections, where conventional antibiotics have failed in this race. However, the development of short cationic antimicrobial peptides with higher efficacy and low cytotoxicity has encouraged our recent exploration. Herein, we describe the synthesis, characterization, and antibacterial evaluation of tetrahydropiperic acid (THPA) conjugate αβ-hybrid peptides, THPA-Lys-tBu-β3,3Ac6c-PEA, P1; THPA-Orn-tBu-β3,3Ac6c-PEA, P2, and THPA-Arg-tBu-β3,3Ac6c-PEA, P3. Our investigation revealed peptide P3 exhibited low hemolytic and best safety index along with higher bactericidal potency against MRSA. Combinatorial study with vancomycin suggested synergistic effect. Mechanistic investigations revealed membrane disruption of MRSA by the peptide. This study suggested that peptide P3 could be an effective therapeutic option to resist the emergence of MRSA-related infections.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to the full article PDF.

USD 39.95

Prices may be subject to local taxes which are calculated during checkout

Fig. 1
Fig. 2
Scheme 1
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data availability

Plotted and analysed data in support of the research article were included along with supporting information. The raw data produced throughout the present investigation are accessible from the corresponding author upon reasonable request.

References

  1. Ferrara F, Castagna T, Pantolini B, Campanardi MC, Roperti M, Grotto A, et al. The challenge of antimicrobial resistance (AMR): Current status and future prospects. Naunyn-Schmiedeberg’s Arch Pharm. 2024;397:9603–15.

    Article  CAS  Google Scholar 

  2. Ahmed SK, Hussein S, Qurbani K, Ibrahim RH, Fareeq A, Mahmood KA, Mohamed MG. Antimicrobial resistance: impacts, challenges, and future prospects. J Med, Surg, Public Health. 2024;2:100081.

    Article  Google Scholar 

  3. Algammal AM, Hetta HF, Elkelish A, Alkhalifah D, Hozzein WN, Batiha GE, El Nahhas N, Mabrok MA. Methicillin-Resistant Staphylococcus aureus (MRSA): one health perspective approach to the bacterium epidemiology, virulence factors, antibiotic-resistance, and zoonotic impact. Infect Drug Resistance. 2020;13:3255–65.

    Article  CAS  Google Scholar 

  4. Robinson DA, Enright MC. Evolutionary models of the emergence of methicillin-resistant Staphylococcus aureus. Antimicrobial Agents Chemother. 2003;47:3926–34.

    Article  CAS  Google Scholar 

  5. Savitskaya A, Masso-Silva J, Haddaoui I, Enany S. Exploring the arsenal of antimicrobial peptides: Mechanisms, diversity, and applications. Biochimie. 2023;214:216–27.

    Article  CAS  PubMed  Google Scholar 

  6. Wang L, Wang N, Zhang W, Cheng X, Yan Z, Shao G, Wang X, Wang R, Fu C. Therapeutic peptides: current applications and future directions. Signal Transduct Target Ther. 2022;7:48.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Drayton M, Kizhakkedathu JN, Straus SK. Towards robust delivery of antimicrobial peptides to combat bacterial resistance. Molecules. 2020;25:3048.

  8. Mahlapuu M, Håkansson J, Ringstad L, Björn C. Antimicrobial peptides: an emerging category of therapeutic agents. Front Cell Infect Microbiol. 2016;6:194.

    Article  PubMed  PubMed Central  Google Scholar 

  9. Ma C, He N, Ou Y, Feng W. Design and synthesis of new vancomycin derivatives. ChemistrySelect. 2020;5:6670–3.

    Article  CAS  Google Scholar 

  10. Wu Z-C, Isley NA, Boger DL. N-Terminus alkylation of vancomycin: Ligand binding affinity, antimicrobial activity, and site-specific nature of quaternary trimethylammonium salt modification. ACS Infect Dis. 2018;4:1468–74.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Velkov T, Thompson PE, Nation RL, Li J. Structure− activity relationships of polymyxin antibiotics. J Med Chem. 2010;53:1898–916.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Wang M, Rakesh KP, Leng J, Fang WY, Ravindar L, Channe Gowda D, Qin HL. Amino acids/peptides conjugated heterocycles: a tool for the recent development of novel therapeutic agents. Bioorg Chem. 2018;76:113–29.

    Article  CAS  PubMed  Google Scholar 

  13. De Zoysa GH, Cameron AJ, Hegde VV, Raghothama S, Sarojini V. Antimicrobial peptides with potential for biofilm eradication: synthesis and structure activity relationship studies of battacin peptides. J Med Chem. 2015;58:625–39.

    Article  PubMed  Google Scholar 

  14. Wani NA, Singh G, Shankar S, Sharma A, Katoch M, Rai R. Short hybrid peptides incorporating β-and γ-amino acids as antimicrobial agents. Peptides. 2017;97:46–53.

    Article  CAS  PubMed  Google Scholar 

  15. Kumar D, Rahman Sarkar A, Iqbal Andrabi N, Assim Haq S, Ahmed M, Kumar Shukla S, Ahmed Z, Rai R. Synthesis, characterization, and anti-inflammatory activity of tetrahydropiperine, piperic acid, and tetrahydropiperic acid via down regulation of NF-κB pathway. Cytokine. 2024;178:156578.

    Article  CAS  PubMed  Google Scholar 

  16. Ur Rahim J, Singh G, Shankar S, Katoch M, Rai R. Tetrahydropiperic acid (THPA) conjugated cationic hybrid dipeptides as antimicrobial agents. J Antibiotics. 2021;74:480–3.

    Article  CAS  Google Scholar 

  17. Humphries R, Bobenchik AM, Hindler JA, Schuetz AN. Overview of changes to the clinical and laboratory standards institute performance standards for antimicrobial susceptibility testing, M100. J Clin Microbiol. 2021;59:10–1128.

  18. Lambert R, Pearson J. Susceptibility testing: accurate and reproducible minimum inhibitory concentration (MIC) and non-inhibitory concentration (NIC) values. J Appl Microbiol. 2000;88:784–90.

    Article  CAS  PubMed  Google Scholar 

  19. Pandey BK, Ahmad A, Asthana N, Azmi S, Srivastava RM, Srivastava S, Verma R, Vishwakarma AL, Ghosh JK. Cell-selective lysis by novel analogues of melittin against human red blood cells and Escherichia coli. Biochemistry. 2010;49:7920–9.

    Article  CAS  PubMed  Google Scholar 

  20. Chowdhary R, Rathore A, Sarkar AR, Kumari J, Manhas R, Firdous S, Mahapa A, Rai R. Antibacterial activity of 2-(4-aminopiperidin-4-yl) acetic acid (β3, 3-Pip) and Its peptide conjugated with lauric acid through the side chain against methicillin-resistant Staphylococcus aureus (MRSA). Micro Pathogenesis. 2025;205:107693.

    Article  CAS  Google Scholar 

  21. Upadhyay A, Pal D, Kumar A. Development of an innovative method of Salmonella Typhi biofilm quantification using Tetrahydrofuran and Response Surface Methodology. Micro Pathogenesis. 2025;208:107992.

    Article  CAS  Google Scholar 

  22. Syal K. Novel method for quantitative estimation of biofilms. Curr Microbiol. 2017;74:1194–9.

    Article  CAS  PubMed  Google Scholar 

  23. Manhas R, Rathore A, Havelikar U, Mahajan S, Gandhi SG, Mahapa A. Uncovering the potentiality of quinazoline derivatives against Pseudomonas aeruginosa with antimicrobial synergy and SAR analysis. J Antibiotics. 2024;77:365–81.

    Article  CAS  Google Scholar 

  24. Hall M, Middleton R, Westmacott D. The fractional inhibitory concentration (FIC) index as a measure of synergy. J Antimicrobial Chemother. 1983;11:427–33.

    Article  CAS  Google Scholar 

  25. Firdous S, Sarkar AR, Manhas R, Chowdhary R, Rathore A, Kumari J, Rai R, Mahapa A. Synthesis, characterization, and antimicrobial activity of urea-containing α/β hybrid peptides against Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus. ACS Omega. 2025;10:2102–15.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Huo S, Chen C, Lyu Z, Zhang S, Wang Y, Nie B, Yue B. Overcoming planktonic and intracellular Staphylococcus aureus-associated infection with a cell-penetrating peptide-conjugated antimicrobial peptide. ACS Infect Dis. 2020;6:3147–62.

    Article  CAS  PubMed  Google Scholar 

  27. Oyama LB, Olleik H, Teixeira A, Guidini MM, Pickup JA, Hui B, Vidal N, Cookson AR, Vallin H, Wilkinson T, Bazzolli D, Richards J, Wootton M, Mikut R, Hilpert K, Maresca M, Perrier J, Hess M, Mantovani HC, Fernandez-Fuentes N, Creevey CJ, Huws SA. In silico identification of two peptides with antibacterial activity against multidrug-resistant Staphylococcus aureus. npj Biofilms Microbiomes. 2022;8:58.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Sarkar AR, Kumari J, Rathore A, Chowdhary R, Manhas R, Firdous S, Mahapa A, Rai R. Antimicrobial activity of α/β hybrid peptides incorporating tBu-β3, 3Ac6c against methicillin-resistant Staphylococcus aureus. J Antibiotics. 2024;77:794–801.

    Article  CAS  Google Scholar 

  29. Shankar S, Jyothi D, Rahim J, Pal PC, Singh UP, Rai R. Conformation of achiral α/β hybrid peptides containing glycine and 1-aminocyclohexaneacetic acid. ChemistrySelect. 2022;7:e202104453.

  30. Ur Rahim J, Ahmad SM, Amin T, Chowdhary R, Goswami A, Rai R. Synthesis, conformation and cytotoxic activity of short hybrid peptides containing conformationally constrained 1-(aminomethyl) cyclohexanecarboxylic acid and gabapentin. Peptides. 2022;158:170897.

    Article  CAS  PubMed  Google Scholar 

  31. Xiong J, Ashraf U, Ye J, Cao S. Extracellular vesicles in pathogenic infection, transmission, and immunity. Engineering. 2024;43:228–40.

    Article  CAS  Google Scholar 

  32. Álvarez A, Fernández L, Gutiérrez D, Iglesias B, Rodríguez A, García P. Methicillin-resistant Staphylococcus aureus in hospitals: Latest trends and treatments based on bacteriophages. J Clin Microbiol. 2019;57:10–1128.

  33. Guo X, An Y, Yan T, Jia Y, Jiao R, Cai X, Yang W, Bao G, Sun W, Yang W, Lu N, Xie J. Halogenated sulfono-γ-AApeptides modified cationic AMPs have enhanced stability and therapeutic potential against clinically important MDR infections. ACS Infect Dis. 2025;11:2018–36.

    Article  CAS  PubMed  Google Scholar 

  34. Chen N, Jiang C. Antimicrobial peptides: structure, mechanism, and modification. Eur J Med Chem. 2023;255:115377.

    Article  CAS  PubMed  Google Scholar 

  35. Sharif S, Yadav AK. Bacterial biofilm and its role in antibiotic resistance. Microbe. 2025;7:100356.

    Article  Google Scholar 

Download references

Acknowledgements

RR acknowledges support by the research project GAP-3102 under the NDTL initiative. AM acknowledges Indian Council of Medical Research, India (ICMR) Research Grant IIRP2023-0715, Council of Scientific and Industrial Research (CSIR) CSIR-MMP075202 and ANRF-SRG SRG/2023/000145 projects for financial support. The authors thank the CSIR-Indian Institute of Integrative Medicine (CSIR-IIIM), Jammu, for providing the necessary environment, facilities, and instrumentation. AR, JK, and SF thank the Council of Scientific and Industrial Research (CSIR), India, and the University Grant Commission (UGC), India, for providing the fellowship grants. RM thanks the Indian Council of Medical Research (ICMR) for providing the project fellowship. The authors acknowledge Dr. Ramajayan Pandian for providing the Rabbit’s blood sample for the hemolysis assay (Institutional Animal Ethics Committee IAEC No.406/85/8/2024). The manuscript have assigned an institutional communication no. CSIR-IIIM/IPR/00957.

Author information

Author notes

  1. These authors contributed equally: Arti Rathore, Beenish Rashid

Authors and Affiliations

  1. Infectious Diseases Division, CSIR-Indian Institute of Integrative Medicine, Jammu, Jammu and Kashmir, India

    Arti Rathore, Shifa Firdous, Rakshit Manhas, Biplab Sarkar, Jyoti Kumari & Avisek Mahapa

  2. Academy of Scientific & Innovative Research (AcSIR), Ghaziabad, India

    Arti Rathore, Aminur Rahman Sarkar, Shifa Firdous, Rubina Chowdhary, Rakshit Manhas, Jyoti Kumari, Rajkishor Rai & Avisek Mahapa

  3. Quality Management & Instrumentation Division, CSIR-Indian Institute of Integrative Medicine, Jammu, Jammu and Kashmir, India

    Beenish Rashid, Aminur Rahman Sarkar, Rubina Chowdhary, Faizan Ahmad Magray & Rajkishor Rai

Authors
  1. Arti Rathore
    View author publications

    Search author on:PubMed Google Scholar

  2. Beenish Rashid
    View author publications

    Search author on:PubMed Google Scholar

  3. Aminur Rahman Sarkar
    View author publications

    Search author on:PubMed Google Scholar

  4. Shifa Firdous
    View author publications

    Search author on:PubMed Google Scholar

  5. Rubina Chowdhary
    View author publications

    Search author on:PubMed Google Scholar

  6. Rakshit Manhas
    View author publications

    Search author on:PubMed Google Scholar

  7. Faizan Ahmad Magray
    View author publications

    Search author on:PubMed Google Scholar

  8. Biplab Sarkar
    View author publications

    Search author on:PubMed Google Scholar

  9. Jyoti Kumari
    View author publications

    Search author on:PubMed Google Scholar

  10. Rajkishor Rai
    View author publications

    Search author on:PubMed Google Scholar

  11. Avisek Mahapa
    View author publications

    Search author on:PubMed Google Scholar

Contributions

AR and BR contributed correspondingly to the research. AR and BR accomplished the experiments and analyzed the whole data. ARS, JK, RM, BS and SF aided in experimentation, data collection, analysis, and interpretation. RC and FM provided additional support in experimental execution and helped with preliminary data processing. RR and AM contributed to conceptualization, methodology expansion, overall guidance, critical revision of the manuscript and supervision. RR and AM equally supervised the whole research. All authors in the article read and accepted the final manuscript.

Corresponding authors

Correspondence to Rajkishor Rai or Avisek Mahapa.

Ethics declarations

Conflict of interest

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Rathore, A., Rashid, B., Sarkar, A.R. et al. Antibacterial activity and mechanism of optimized THPA conjugated dipeptides against methicillin-resistant Staphylococcus aureus. J Antibiot 79, 15–29 (2026). https://doi.org/10.1038/s41429-025-00877-w

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Version of record:

  • Issue date:

  • DOI: https://doi.org/10.1038/s41429-025-00877-w

Search

Advanced search

Quick links