Electron and Proton Transfers Modulate DNA Binding by the Transcription Regulator RsrR

Abstract

The [Fe2S2]-RsrR gene transcription regulator senses the redox status in bacteria by modulating DNA binding while its cluster cycles between +1 and +2 states-only the latter binds DNA. We have previously shown that RsrR can undergo remarkable conformational changes involving a 100° rotation of tryptophan 9 between exposed (Out) and buried (In) states. Here, we have used the chemical modification of Trp9, site-directed mutagenesis and crystallographic and computational chemical studies to show that (i) the Out and In states correspond to oxidized and reduced RsrR, respectively, (ii) His33 is protonated in the In state due to a change in its pKa caused by cluster reduction and (iii) Trp9 rotation is conditioned by the response of its dipole moment to environmental electrostatic changes. Our findings illustrate a novel function of protonation resulting from electron transfer.

Identifying Free Energy Landscapes of Proton-Transfer Processes between Brønsted Acid Sites and Water Clusters Inside the Zeolite Pores

“…AIMD simulations combined with the enhanced sampling technology is one of the most powerful methods in the arsenal of computational chemistry to examine the movements overcoming pronounced high barriers in the entire free energy landscape. − This method has been applied to successfully elucidate the proton hopping mechanism within water clusters, − proteins, − metal oxides, − and polymer membranes . In the present work, the proton-transfer processes between the BAS site and water molecules inside the MFI zeolite pores were investigated for the first time.…”

How Oxidation of a Unique Iron-Sulfur Cluster in FBXL5 Regulates IRP2 Levels and Promotes Regulation of Iron Metabolism Proteins

“…Although the sensing mechanism of ISCs has thus far been reported to depend on changes in the stoichiometry or integrity of the cluster, including its partial or total disassembly (as in FNR or IRP1), it has remained unclear how a change in the oxidation state of the cluster itself is capable of affecting the affinity of FBXL5 binding to IRP2. A recent study on the bacterial sensor RsrR reported that the transition of its [2Fe-2S] cluster from +1 to +2 overall charge was associated with a large change in the RsrR protein conformation, which in turn determined its ability to bind transcriptional binding sites and reprogram expression of target genes (Crack et al, 2020). It is therefore conceivable that a similar conformational rearrangement may accompany changes in the oxidation state of the [2Fe-2S] cluster of FBXL5, thus explaining its increased binding to IRP2 upon oxidation.…”

Transient Formation of a Second Active Site Cavity during Quinolinic Acid Synthesis by NadA

“…In the Schrödinger suite, by default and for any force field, the charges of [Fe-S] clusters are formal, that is, +2 or +3 for the Fe atoms and −2 for the inorganic bridging S atoms. We know from previous MD studies on the RsrR [FeS]-containing metalloprotein that these formal charges can induce artifactual interactions. Accordingly, we calculated more realistic charges using a small quantum model (Figure S10).…”

“…31 Figure 3 and Figure S10 were prepared with Maestro. 36 MD Simulations. Calculations were performed with programs from the Schrodinger suite.…”

“…Calculations were performed with programs from the Schrodinger suite. 36 Because we wanted to investigate the rotation of Phe58 in an open form of TmNadA, where the formation of cavity II is not possible (see main text), the new TmNadA*Y107F crystal structure was used. It is at a higher resolution than the open TmNadA* (pdb code 4p3x).…”