Nocardia are opportunistic human pathogens that can cause a range of debilitating and difficult to treat infections of the lungs, brain, skin, and soft tissues. Despite their close relationship to the well-known secondary metabolite-producing genus, Streptomyces, comparatively few natural products are known from the Nocardia. Here, we combine chemistry, genomics, and molecular microbiology in conjunction with an 8,500 member transposon mutant library to link genes to new molecules in human pathogenic Nocardia. We have used this library to unveil the biosynthetic pathway to a new cytotoxic and antifungal polyene, terpenomycin, in a human pathogenic Nocardia isolate; to identify the basis for antibacterial activity in this same isolate and to induce the production of new secondary metabolites in this strain. Our findings show that the neglected actinomycetes, such as the Nocardia, have an unappreciated capacity for the production of bioactive molecules with unique biosynthetic pathways waiting to be uncovered and highlights these organisms as producers of diverse natural products.