Introduction: Antimicrobial peptides can be used as an antimicrobial agent against a wide range of microorganisms. Thanatin is a 21 amino acid insect-derived antimicrobial peptide (GSKKPVPIIYCNRRTGKCQRM). It was shown that E. coli, Salmonella typhimurium, Klebsiella pneumoniae, and Staphylococcus aureus were completely sensitive to this peptide while Listeria monocytegenes and Pseudomonas aeruginosa were more resistant to thanatin (Javadmanesh et al, 2018 and Tanhaeian et al, 2019).
This peptide has the potential to be used as an antibiotic substitute. Accordingly, thermo-stability of these antimicrobial agents at different temperatures for processing purposes could be considered as an advantage. In this study, we predicted the thermal-stability of thanatin in different thermal circumstances through in silico analysis.
Methods: Thermal stability
Vulnerability of thanatin against high temperatures was estimated under different time and thermal circumstances by Molecular Dynamic (MD) simulations. Three different conditions were considered according to the probable agricultural use of therapeutic peptides. Accordingly, normal dairy cattle body temperature (312 º K), normal avian body temperature (315 º K), and water boiling temperature at sea level were applied.
Dynamic simulation
In order to predict the thermal-susceptibility of thanatin, the crystallography structure (PDB entry: 8TFV) of this peptide was used. GROMACS 5.0.1 (Maupetit et al. 2010), GROMOS 54a7 (Karplus et al. 2002) force field, SPC/E water molecules (Narayanan et al. 2007), and cubic box with periodic boundary conditions were utilized for MD simulation. 100 ns and 200 ns MD simulation were performed for normal body temperatures and water boiling temperature at sea level, respectively. The root-mean-square deviation (RMSD) of thanatin α-carbons was plotted versus time during the 100 and 200 ns MD simulation.
Results: The RMSD results for three candidate temperatures illustrated the thermo-stability of thanatin at all three conditions. Despite steady RMSD of thanatin in dairy cattle and avian normal body temperatures, the fluctuations of RMSD for thanatin in water boiling temperature depict that this peptide was not stable during the first 150 ns. After a stabilization between 100ns and 150ns which showed temporary stability, the RMSD fell considerably and the value of RMSD remained steady until the end of simulations with an approximate 0.1 nm deviation.
The tertiary structure of thanatin changed significantly during these three MD simulations. The more we escalate temperature the more tertiary structure changed as the conformation of thanatin was denatured in water boiling temperature.
Conclusion: Thanatin is stable in high temperature and it might be possible to use it as an antibiotic substitute or bio-preservative in food processing where heat treatment is a part of processing. Further experiments are required to prove this concept.
References
Javadmanesh, A., Tanhaeian, A., Mousavi, S.Z., Azghandi, M. 2018. Investigation of recombinant thanatin effects on the growth inhibition of e. coli mastitis in dairy cows. Proceedings of the 2nd International Congress on Biomedicine. Tehran, Iran. 24-26 Dec, 2018.
Karplus, M., McCammon, J. A. (2002). Molecular dynamics simulations of biomolecules. Nat. Struct. Mol. Biol, 9(9), 646.
Maupetit, J., Derreumaux, P., Tufféry, P. (2010). A fast method for large‐scale De Novo peptide and miniprotein structure prediction. J. Comput. Chem, 31(4), 726-738.
Narayanan, R., & Johnston, D. (2007). Long-term potentiation in rat hippocampal neurons is accompanied by spatially widespread changes in intrinsic oscillatory dynamics and excitability. Neuron, 56(6), 1061-1075.
Tanhaeian, A., Mousavi, S.Z., Azghandi, M., Javadmanesh, A. 2019. Expression of Thanatin in HEK293 Cells and Investigation of Its Antibacterial Effects on Some Human Pathogens. Protein and Peptide Letters. 26: doi: 10.2174/0929866526666190822162140.