Date Published: July 25, 2018
Author(s): Harkirat S. Sethi, Jessica L. Osier, Geordan L. Burks, Jennifer F. Lamar, Hana McFeeters, Robert L. McFeeters.
New antibiotics and new antibiotic targets are needed to counter the development of bacterial drug resistance that threatens to return the human population to the pre-antibiotic era. Bacterial peptidyl-tRNA hydrolase (Pth1) is a promising new antibiotic target in the early stages of development. While inhibitory activity has been observed in a variety of natural products, bioactive fractionation has been a bottleneck for inhibitor isolation. To expedite the isolation of inhibitory compounds from complex mixtures, we constructed a Pth1 affinity column and used it to isolate inhibitory compounds from crude natural products. Recombinantly produced S. typhimurium Pth1 was covalently attached to a column matrix and the inhibitory activity isolated from ethanol extracts of Salvinia minima. The procedure reported here demonstrates that isolation of Pth1 inhibitory compounds from complex natural product extracts can be greatly expedited over traditional bioactive fractionation, decreasing time and expense. The approach is generally applicable to Pth1s from other bacterial species and opens an avenue to advance and accelerate inhibitor development against this promising antimicrobial target.
Antibiotic resistance has become a significant problem in both public health and the field of medicine. As the use of antibiotics has increased, bacteria have developed tolerance or become resistant. Some bacteria, such as certain strains of Salmonella typhimurium, have become multidrug resistant [1,2]. As a consequence, drug treatments have become less effective and, in the United States, at least 2 million people get bacterial infections and at least 23,000 die annually . Invasive Salmonella disease remains a major health problem, particularly in many parts of Asia and Africa . To counter the rise of antibiotic resistance, in Salmonella spp and other pathogenic bacteria, new antibiotics and new antibiotic targets are urgently needed.
As drug discovery targeting the promising new antibiotic target Pth1 advances, the bottleneck of inhibitor isolation remains a significant barrier. Currently screening throughput is limited due to characterization of Pth1 functional characterization. Potential exists for increasing throughput, including development of higher throughput fluorescence polarization methods . However, significant barriers exist for scale up of this methodology. Options like fragment based drug design are desirable, but again are limited by functional characterization.