C. diphtheriae toxin

The diphtheria toxin repressor contains an SH3-like domain that forms an intramolecular complex with a proline-rich (Pr) peptide segment and stabilizes the inactive state of the repressor. Upon activation of diphtheria toxin repressor (DtxR) by transition metals, this intramolecular complex must dissociate as the SH3 domain and Pr segment form different interactions in the active repressor. Here we investigate the dynamics of this intramolecular complex using backbone amide nuclear spin relaxation rates determined using NMR spectroscopy and molecular dynamics trajectories. The SH3 domain in the unbound and bound states showed typical dynamics in that the secondary structures were fairly ordered with high generalized order parameters and low effective correlation times, while residues in the loops connecting beta-strands exhibited reduced generalized order parameters and required additional motional terms to adequately model the relaxation rates. Residues forming the Pr segment exhibited low-order parameters with internal rotational correlation times on the order of 0.6 ns(-1) ns. Further analysis showed that the SH3 domain was rich in millisecond time scale motions while the Pr segment exhibited motions on the 100 mu s time scale. Molecular dynamics simulations indicated structural rearrangements that may contribute to the observed relaxation rates and, together with the observed relaxation rate data, suggested that the Pr segment exhibits a binding < - > unbinding equilibrium. The results here provide new insights into the nature of the intramolecular complex and provide a better understanding of the biological role of the SH3 domain in regulating DtxR activity. (C) 2007 Elsevier Ltd. All rights reserved.

The mycobacterial IdeR protein is a homologue of the diphtheria-toxin repressor DtxR. We have previously demonstrated that Mycobacterium tuberculosis IdeR, like DtxR, represses transcription of Corynebacterium diphtheriae iron-regulated promoters in vivo and binds to C. diphtheriae operators in a metal-dependent manner in vitro. We show here that ideR mutants of M. smegmatis, constructed by allelic replacement, were defective in their ability to repress siderophore biosynthesis in the presence of iron. They were also more sensitive to hydrogen peroxide and had decreased levels of catalase/peroxidase (KatG) and manganese superoxide dismutase (Mn-SOD). This indicates that IdeR is a negative regulator of siderophore production and is required for the response to superoxide- and hydrogen peroxide stress. We propose that IdeR is the mycobacterial counterpart of the Escherichia coli Fur protein, i.e. it is a pleiotropic regulator that couples iron metabolism to the oxidative-stress response.