Premium
MsbA as a Drug Discovery Target for Compounds with Antibacterial Activities
Author(s) -
LaVigne Collette A,
McCormick James W,
Vogel Pia D,
Wise John G
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.939.14
Subject(s) - atp binding cassette transporter , chemistry , antibiotics , bacteria , lipid ii , biochemistry , in vivo , enzyme , in vitro , bacterial outer membrane , drug discovery , escherichia coli , biology , transporter , biosynthesis , microbiology and biotechnology , gene , genetics
There is an ever increasing need for novel antibiotics today as many bacteria adapt and become resistant to commonly used antibiotics. MsbA is a prokaryotic integral membrane protein, one of the ATP‐binding cassette (ABC) transporters, and transports lipid A from the outer to the inner leaflet of the bacterial plasma membrane. MsbA is essential in gram‐negative bacteria for proper membrane formation and might therefore be viewed as an antibiotic target. The fact that MsbA also transports a wide variety of drug‐like compounds across the membrane, however, complicates the search for effective inhibitors. In efforts similar to those described in (Brewer et al. Mol Pharmacol. 2014) for the search for inhibitors of the multidrug resistance protein, P‐glycoprotein, computational screens were performed to search for drug‐like compounds that effectively inhibit MsbA ATP hydrolysis without being themselves transported by it. Identified compounds were purchased and tested in both in vivo and in vitro models to determine efficacy. A His 6 ‐tagged MsbA was overexpressed and purified from E. coli and reconstituted into nanodiscs, where MsbA has been shown in the past to have high catalytic activity. ATP hydrolysis rates of MsbA were measured in the presence and absence of potential inhibitors using a coupled enzyme assay. E. coli growth assays with in the presence of identified compounds were used as an in vivo model to evaluate potential growth inhibition. Synergistic effects were also studied using growth assays where potential inhibitor compounds were administered in addition to sub‐lethal concentrations of known antibiotics. Preliminary results suggest that several compounds identified by the computational screening inhibited MsbA ATP hydrolysis activity. Further experiments will aim to better characterize and optimize these compounds for potential use as novel antibiotics. Support or Funding Information This work is supported by the SMU University Research Council, the SMU Center for Drug Discovery, Design and Delivery, the Communities Foundation of Texas, and a private gift from Ms. Suzy Ruff of Dallas, Texas.