2026-10-12T19:03:00Z https://api.figshare.com/v2/oai

oai:figshare.com:article/32641935 2026-10-01T16:19:35Z portal_1172 item_type_8 month_year_10_2026

Stepwise structural optimisation and functional evaluation of dermaseptin-DI2, an antimicrobial peptide identified from Phyllomedusa tomopterna Lingyu Ke (24164634) PUREID: 659005140 Antimicrobial peptides (AMPs) C-terminal truncation machine learning-guided substitutions hydrocarbon stapling–prodrug integration therapeutic index serum and salt stability A stepwise optimisation framework was developed to enhance the therapeutic potential of antimicrobial peptides (AMPs) against multidrug-resistant (MDR) pathogens, integrating C-terminal truncation to improve therapeutic index (TI), targeted amino acid substitutions to enhance serum stability, and hydrocarbon stapling–prodrug integration to boost salt stability while mitigating host toxicity. In Chapter 3, dermaseptin-DI2, a frog-skin-derived peptide, underwent rational two-step C-terminal truncation, yielding DI2(1–19) with a > 50-fold TI improvement over the parent while retaining potent, broad-spectrum activity and rapid bactericidal kinetics. Both peptides maintained stable MICs over 15 passages, outperforming first-line antibiotics, although DI2(1–19) showed reduced ionic stability and antibiofilm potency. In Chapter 4, DI2(1–19) was further optimised at the proteolytic site Lys⁴ via machine learning-guided substitutions with unnatural Lys analogues. Among these, [Orn⁴]DI2(1–19) and [Dab⁴]DI2(1–19) retained bactericidal speed, inhibited biofilms, and improved activity retention in human serum by up to seven-fold over DI2(1–19). In vivo, both were well tolerated and achieved superior efficacy to polymyxin B (PMB) against P. aeruginosa and vancomycin (VAN) against MRSA, clearing infections within 3 h. In Chapter 5, to address poor salt stability, DI2(1–19) was modified via hydrocarbon stapling, enhancing salt resistance and spectrum but increasing cytotoxicity. The lead stapled analogue V10A14 was further converted into a prodrug (V-KLE) by attaching a neutrophil elastase (NE)-cleavable linker and polyglutamate masking group. V-KLE was inactive and non-toxic until enzyme activation restored full potency, while also significantly improving proteolytic stability. Collectively, this tiered approach generated AMP candidates with high TI, stability across challenging physiological environments, and potent in vivo efficacy, offering a transferable blueprint for next-generation peptide antibiotic design. Thesis is embargoed until 31 December 2030. 2026-10-01T16:19:35Z Text Thesis 10.17034/32641935.v1 https://figshare.com/articles/thesis/Stepwise_structural_optimisation_and_functional_evaluation_of_dermaseptin-DI2_an_antimicrobial_peptide_identified_from_i_Phyllomedusa_tomopterna_i_/32641935 All Rights Reserved Open Access after 2030-12-31