Structure and function of the DNR transcription factor, a master regulator of nitric oxide homeostasis in Pseudomonas aeruginosa

Understanding the fate of nitric oxide (NO) inside the bacterial cell is a major issue in biology of host/pathogen interactions. Denitrifying bacterial species keep intracellular NO below cyto- toxic levels by regulating of the expression and activity of the enzymes involved in its turnover. Expression of these genes is controlled by redox-linked transcription factors of the CRP-FNR superfamily of regulators, such as FNR, DNR and NNR (1,2). These proteins present a dimerization domain, a DNA binding helix-turn-helix domain and a sensor domain. involved in the response to different signal molecules, such as O2, CO and NO. The DNR regulator from Pseudomonas aeruginosa has a master role in the control of NO homeostasis in this opportunistic human pathogen (3). DNR is active under low oxygen tension in the presence of N-oxides: in particular, it has been suggested that it may act as a NO sensor in vivo (4). DNR forms a stable complex with heme (5); this cofactor is required in the bacterium to perceive the physiological messenger, i.e. nitric oxide (6). Here we present a spectroscopic, kinetic and protein engineering study which allowed us to propose the amino- acid residues involved in heme iron coordination in DNR. The results of this study underlines that DNR is characterized by a remarkable flexibility in solution, in agreement with the data avail- able on other members of the CRP-FNR class of transcriptional regulators. We propose that protein flexibility and dynamics is a key structural feature essential to explain the evolutionary success and adaptability of these bacterial transcription factors (7). References: 1. Giardina, G et al. Biochem Soc Trans 2011; 39: 294–298. 2. Giardina, G et al. Proteins 2009; 77: 174–80. 3. Hassett, DJ et al. Trends Microbiol 2009; 17: 130–138. 4. Spiro, S. FEMS Microbiol Rev 2007; 31: 193–211. 5. Giardina, G et al. J Mol Biol 2008; 378: 1002–1015. 6. Castiglione, N et al. Microbiology 2009; 155: 2838–2844. 7) Rinaldo et al. 2011 submitted.