In this study, we have re-evaluated the myxobacterial antibiotic myxovalargin and found it to be bactericidal on several important human and animal pathogens at much lower MICs than described and anticipated from previous studies including tuberculosis-causing bacilli. Through identification of an improved natural producer and its fermentation compound availability was ensured to enable in vivo efficacy and early preclinical studies as reported here. Importantly, by studying the molecular details of Myx biosynthesis two misassignments in the stereochemistry of the reported structure were identified which then provided the basis for the development of a concise synthetic route ultimately verifying the corrected structural assignment. Having the compound in hand also allowed the determination of the ribosome as the molecular target which was found in a combination of genome sequencing of resistant mutants raised in M. tuberculosis and subsequent structural analysis of MyxA binding to the bacterial 70S ribosome. Although the binding site of MyxA on the ribosome overlaps significantly with other antimicrobial peptides, such as Api137 19 , Bac7 , klebsazolicin and phazolicin , the conformation and interaction mode are quite distinct (Fig. ). Unlike other tunnel binding antibiotics, such as Ery and TcmX, the compact conformation of MyxA completely occludes the NPET, in a manner reminiscent of the blockage caused by the synergistic streptogramin dalfopristin and quinupristin combination (Fig. ) . These findings suggest that the mechanism of action of MxyA is also analogous to the streptogramins, namely, to trap ribosomes during a late stage of translation initiation by preventing full accommodation of the intiator tRNA at the peptidyl-transferase center on the large ribosomal subunit. MyxA shows promising pharmacokinetic properties and an in vivo efficacy proof-of-concept was provided in a mouse model of sepsis caused by the bovine pathogen P. multocida. However, toxicity observed when administering MyxA in a more challenging mouse model of tuberculosis is presumably caused by an unspecified "membrane effect" and currently prevents further development towards tuberculosis treatment. Nevertheless, our studies set the stage to overcome this hurdle as improving the safety margin of myxovalargins seems feasible due to three main findings from this work: The molecular understanding of MyxA-ribosome interaction, as well as chemical and biotechnological access to the natural product scaffold should allow to differentiate the specific ribosome inhibitory effect from the unspecific toxicity to enable preclinical development of myxovalargin as a potential antibiotic in the future.