Development of high-throughput screening method for iron transport inhibitors in E. coli

Date

2015-01-14

Journal Title

Journal ISSN

Volume Title

Publisher

Kansas State University

Abstract

Iron acquisition is a component of Gram-negative bacteria pathogenesis, therefore as a form of 'nutritional immunity' host organisms sequester iron. To obtain iron bacteria secrete siderophores that scavenge iron. The E. coli outer membrane protein FepA actively transports the siderophore ferric enterobactin into the periplasm. We observe this uptake reaction by fluorescently labeling FepA in live bacteria, monitoring quenching that occurs upon binding of FeEnt, and then fluorescence recovery during transport. Energy poisons azide, arsenate, and 2,4-dinitrophenol were evaluated to determine sensitivity to known transport inhibitors. We developed and optimized methods to screen for iron transport inhibitors using a cell-based high-throughput screening platform. These inhibitors may have broad spectrum bacteriostatic antibiotic properties.

Description

Keywords

Iron, FepA, TonB, High-throughput screening, Biochemistry

Graduation Month

May

Degree

Master of Science

Department

Department of Biochemistry and Molecular Biophysics

Major Professor

Phillip E. Klebba

Date

2015

Type

Thesis

Citation