Antimicrobial peptides (AMPs) like tritrpticins, exhibit non-specific membrane lysis of gram-negative bacteria and can replace antibiotics, combating multi-drug resistance observed in UTI patients. Tritrpticins designated–NT, T1, T2, T3, T5, T7 and T8, were computationally investigated by interaction with Escherichia coli membrane model, mammalian cell toxicity and structural stability to identify a potential drug scaffold for UTI. Initially T3 was eliminated due to low interaction with Escherichia coli membrane model, based on its computed solvation energy. Further, negative support vector machine (SVM) scores revealed non-toxicity of T1, T2, T5, T7 and T8. Finally, at 310 K and varying pH 4.5–9.0, T5 exhibited highest structural stability based on its highest consistency of hydrogen bonds (H-bonds), root mean square deviation (RMSD) and secondary structure profiles along with its lowest conformational free energy. Overall, T5 could be considered a promising peptide drug scaffold to combat UTI. ABBREVIATIONSAMP antimicrobial peptidePBEQ Poisson Boltzmann equationH-bonds hydrogen bondsMIC minimum inhibitory concentrationLD50 lethal dose, 50%RMSD root mean square deviationSVM support vector machineUTI urinary tract infection Communicated by Ramaswamy H. Sarma. © 2019 Informa UK Limited, trading as Taylor & Francis Group.