Alternating Pattern of Stereochemistry in the Nonactin Macrocycle Is Required for Antibacterial Activity and Efficient Ion Binding

Nonactin is a polyketide antibiotic produced by Streptomyces griseus ETH A7796 and is an ionophore that is selective for K+ ions. It is a cyclic tetraester generated from two monomers of (+)-nonactic acid and two of (−)-nonactic acid, arranged (+)-(−)-(+)-(−) so that nonactin has S4 symmetry and is achiral. To understand why achiral nonactin is the naturally generated diastereoisomer, we generated two alternate diastereoisomers of nonactin, one prepared solely from (+)-nonactic acid and one prepared solely from (−)-nonactic acid, referred to here as ‘all-(+)-nonactin’ and ‘all-(−)-nonactin’, respectively. Both non-natural diastereoisomers were 500-fold less active against Gram positive organisms than nonactin confirming that the natural stereochemistry is necessary for biological activity. We used isothermal calorimetry to obtain the K a, ΔG, ΔH, and ΔS of formation for the K+, Na+, and NH4 + complexes of nonactin and all-(−)-nonactin; the natural diastereoisomer bound K+ 880-fold better than all-(−)-nonactin. A picrate partitioning assay confirmed that all-(−)-nonactin, unlike nonactin, could not partition K+ ions into organic solvent. To complement the thermodynamic data we used a simple model system to show that K+ transport was facilitated by nonactin but not by all-(−)-nonactin. Modeling of the K+ complexes of nonactin and all-(−)-nonactin suggested that poor steric interactions in the latter complex precluded tight binding to K+. Overall, the data show that both enantiomers of nonactic acid are needed for the formation of a nonactin diastereoisomer that can act as an ionophore and has antibacterial activity.