Affinity selection-mass spectrometry identifies a novel antibacterial RNA polymerase inhibitor.

Walker, SS, Degen D, Nickbarg E, Carr D, Soriano A, Mandal M B, Painter RE, Sheth PR, Xiao L, Sher X, Murgolo N, Su J, Olsen DB, Ebright RH, Young K.  2017.  

Journal:

ACS Chemical Biology

Abstract:

The growing prevalence of drug-resistant Gram-negative bacteria is a significant global threat to human health. Rifampicin, an RNA polymerase-targeting agent, is an important part of the antibacterial armamentarium; however the emergence of resistance requires that it be used against only certain infections and usually in combination with another antibiotic. While rifampicin has significant clinical limitations, it does show that bacterial RNA polymerase can be an effective target for antibacterial intervention. To find new RNA polymerase inhibitors we initiated a screen of a collection of antibacterial bioactive molecules using affinity selection-mass spectrometry and purified Escherichia coli core RNA polymerase (subunits α, β, β', ω). Affinity selection screening identified a novel small molecule, MRL-436, that binds selectively to and inhibits RNA polymerase in vitro and inhibits RNA synthesis in the cell. Selection for resistance followed by whole genome sequencing identified a missense mutation in rpoC (β' subunit) and, separately, frameshift mutations in rpoZ (ω subunit) suggesting that MRL-436 targets RNA polymerase in the cell. In addition, cells lacking the rpoZ gene or purified RNA polymerase containing either a specific substitution in β' or lacking ω are selectively resistant to MRL-436. Molecular modeling indicates that the location of the substitution mutation in β' is closely juxtaposed with ω in a region of the complex thought to be important for transcription regulation during cellular response to amino acid starvation.

Citation:
Walker, SS, Degen D, Nickbarg E, Carr D, Soriano A, Mandal M B, Painter RE, Sheth PR, Xiao L, Sher X et al..  2017.  Affinity selection-mass spectrometry identifies a novel antibacterial RNA polymerase inhibitor.. ACS Chemical Biology.