Metal Ion Binding of The Corynebacterium pseudotuberculosis Diphtheria Toxin Repressor
Journal Title: Biomedical Journal of Scientific & Technical Research (BJSTR) - Year 2019, Vol 14, Issue 2
Abstract
The acquisition of iron is essential to facilitate growth of bacterial pathogens. Bacteria using high-affinity iron uptake systems, siderophores, to scavenge iron from the host is regulated by the diphtheria toxin repressor (DtxR). Additionally, DtxR is a global regulator of cell metabolism and regulates the expression of diphtheria toxin in C. diphtheriae. The genome of Corynebacterium pseudotuberculosis, a related animal pathogen with C. diphtheriae, contain the genes for diphtheria toxin and DtxR. Our study describe the high scale heterologue expression of Corynebacterium pseudotuberculosis DtxR (Cp-DtxR) in E.coli, purification, and characterization of the Fe2+ binding using CD spectroscopy.The acquisition of essential nutrients and ions such as iron is of primordial significance for the growth and development of bacterial pathogens [1]. Since the concentration of free iron in human serum is estimated to be 10-12 μM, which is much lower than that required for optimal bacterial growth [2], invasive pathogenic bacteria overcome this problem by using inducible, high-affinity iron uptake systems, siderophores, to scavenge iron from the host [3,4]. In gram-positive and gram-negative bacteria, the diphtheriae toxin repressor (DtxR) plays a key role in the regulation and production of siderophores and genes involved in the uptake of iron and other transition metals [5-10]. Additionally, DtxR regulates the expression of diphtheriae toxin in C. diphtheriae [11]. The threedimensional structure of C. diphtheriae DtxR (Cd-DtxR) reveals that it exists as a dimer containing two metal binding sites per monomer [12,13]. Several transition metal ions, such as Fe(II), Ni(II), Co(II), Cd(II), Mn(II) and, to some extent, Zn(II) activate apo-Cd-DtxR and, the holo-enzyme binds its target DNA and blocks the transcription of the downstream genes [14-17]. Yellaboina et al. [6] predicted more than 70 DtxR-regulated operons in the C. diphtheriae genome and demonstrated that DtxR is a global regulator of cell metabolism. Our study was performed on DtxR of Corynebacterium pseudotuberculosis, a related pathogen of Corynebacterium diphtheriae. Analysis of the C. pseudotuberculosis genome identified diphtheriae toxin and DtxR genes. However, diphtheriae toxin is not involved in the C. pseudotuberculosis infection. We assume that DtxR in C. pseudotuberculosis mainly plays a role in divalent ion uptake and in the regulation of metabolism. C. pseudotuberculosis causes caseous lymphadenitis (CLA) in equids, sheep, goats, and to a lesser extent in horses and cattle. This disease leads to considerable economic loss in many countries, including Brazil [18,19] and presently, no effective treatment is available to combat this disease. Based on its key role in pathogen cell metabolism, Cp-DtxR can be considered as a potential drug target and we present results of the expression, purification and characterization of Cp-DtxR and the iron binding by Cp-DtxR apo-protein.In Silico Analysis: Cp-DtxR and Cd-DtxR sequences were retrieved from NCBI and a sequence alignment were performed using MUSCLE [20] and Box Shade web servers. The atomic coordinates from C. diphtheriae DtxR (PDB code: 1G3S) were used as the template for comparative modeling by the satisfaction of spatial restraints as implemented in the program Modeller 9v13 [21]. The structure of native Cp-DtxR (Gene ID: ADL10687.1; Uniprot: D9QAW1) was modeled.
Authors and Affiliations
Carolina Gismene, Angela N Santisteban, Monika A Coronado, Raphael J Eberle, Raghuvir K Arni
Keywords
Related Articles
- EP ID EP598533
- DOI 10.26717/BJSTR.2019.14.002513
- Views 156
- Downloads 0
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