Liquid-Phase Peptide Synthesis of Antimalarial Kozupeptins Using a Benzoyl-Type Soluble Hydrophobic Auxiliary

Liquid-phase peptide synthesis (LPPS) has emerged as a powerful platform for constructing complex peptide natural products, yet its efficiency remains highly dependent on carrier design, protecting group compatibility, and late-stage functional group manipulations. Here, we report a streamlined LPPS strategy enabled by a benzoyl-type tag carrier that facilitated tert-butoxycarbonyl (Boc)-based elongation, mild carrier cleavage, and direct access to C-terminal peptide alcohols. This platform enabled the concise synthesis of kozupeptin aldehyde-an exceptionally potent antimalarial peptide aldehyde-in only 11 purification steps from commercially available methyl gallate, representing a substantial improvement over previous methodologies. The robustness of the benzoyl tag system further allowed the rapid preparation of 10 analogs by parallel one-pot LPPS via single-residue scanning across the peptidic core. Biological evaluation against Plasmodium falciparum revealed tight structure-activity relationships, identifying strict sequence and conformational requirements for potency while highlighting the crucial stereoelectronic influence of the 4-methyl-proline (Pro(4-Me)) residue and the threonine (Thr)-proline (Pro) amide bond equilibrium. Collectively, this work establishes a versatile LPPS approach for accelerated access to peptide natural products and provides new insights into the conformational determinants underlying kozupeptin's antimalarial activity.