Iron Superoxide Dismutase is Upregulated in Response to Antibiotic Challenge in Multi‐Drug Resistant Acinetobacter baumannii

Abstract

Acinetobacter baumannii is an opportunistic gram‐negative human pathogen. Underscoring the threat to global health is the observation of isolates of A. baumannii that are resistant to all known antibiotics. Multidrug resistant A. baumannii is a leading cause of nosocomial infections and can lead to increased mortality and length of stay. To address this public health problem, there is a need to develop new therapeutics. Our approach is to identify potential drug targets by characterizing molecular mechanisms that contribute to the antibiotic resistance of A. baumannii. To achieve this, we leverage a collated bank of 247 isolates (72% are multi‐drug resistant) of A. baumannii from patients at Nashville General Hospital. Isolate MMC4 (Meharry Medical College Isolate 4) is resistant to >25 antibiotics and hence represents an ideal system for dissecting the molecular origins of antibiotic resistance. Here we report the identification of superoxide dismutase B (SodB) as an important antibiotic resistance factor along with its biochemical characterization and high‐resolution crystal structure. Isolate MMC4 was subjected to proteomic analysis in the presence and absence of antibiotic challenge using multidimensional protein identification technology. SodB showed the greatest differential protein expression, a greater than 40 fold increase in response to antibiotic challenge. SodB is an enzyme that protects bacteria from oxidative stress by dismutating highly reactive superoxide radicals. While the superoxide dismutase family has been extensively studied, little is known about A. baumannii SodB. We expressed SodB recombinantly from E. coli and purified it to >95% homogeneity. We evaluated the activity of SodB using a tetrazolium salt assay and established it is an iron dependent enzyme using inductively coupled plasma – optical emission spectrometry. We also determined the structure of SodB with iron bound to 1.45 angstroms resolution. The crystal belonged to the C121 monoclinic space group with unit cell sides (angstroms) and angles (degrees) 97.28, 40.74, 76.63 and 90, 123, 90, respectively. In total, these data establish the importance of protection from oxidative stress in antibiotic resistant A. baumannii and provide the first steps for a structure‐based drug design approach to target inhibition of SodB as a therapeutic strategy.Support or Funding InformationThis work was funded by National Science Foundation HRD1547757, National Institutes of Health R25MD010396 and R01HD090061, and Office of Medical Research, Department of Veterans Affairs IK2BX001701.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.