Griselimycins - magic bullets against Mycobacterium tuberculosis?

Despite modern antibiotics and the development of a curative regimen for tuberculosis, this disease remains a worldwide problem and the emergence of drug-resistant Mycobacterium tuberculosis has prioritised the need for new drugs. Here we show that new and optimised derivatives from Streptomyces-derived griselimycin are highly active against M. tuberculosis, both in vitro and in vivo. After identification of the griselimycin biosynthetic gene cluster we were able to clarify the biosynthesis of the biosynthetic precursor methyl-proline which is incorporated into the natural product at the site of metabolic lability. This finding opened up oportunities to improve the ratio of metabolically stable versus unstable griselimycin derivatives. Remarkably, methylproline is generated from leucine by a specific pathway involving four enzymes expression of which results in methy-proline formation in heterologous hosts. Based on self-resistance studies in Streptomyces and genomic analyses of resistant mycobacteria, we found that griselimycins inhibit the DNA polymerase sliding clamp DnaN; these interactions were characterised by surface plasmon resonance and crystal structure analysis. Furthermore, we discovered that infrequent resistance to griselimycin is associated with highly unusual target amplification in mycobacteria. Our results demonstrate that griselimycin and its derivatives have high translational potential for tuberculosis, validate DnaN as an antimicrobial target and capture the process of antibiotic pressure-induced target amplification.