Search

Dimethylsulfoniopropionate (DMSP) is an important marine anti-stress compound, with key roles in global nutrient cycling, chemotaxis and, potentially, climate regulation. Recently, diverse marine Actinobacteria, α- and γ-proteobacteria were shown to initiate DMSP synthesis via the methionine (Met) S-methyltransferase enzyme (MmtN), generating S-methyl-Met (SMM). Here we characterize a roseobacterial MmtN, providing structural and mechanistic insights into this DMSP synthesis enzyme. We propose that MmtN uses the proximity and desolvation mechanism for Met S-methylation with two adjacent MmtN monomers comprising the Met binding site. We also identify diverse functional MmtN enzymes in potentially symbiotic archaeal Candidatus Woesearchaeota and Candidate Phyla Radiation (CPR) bacteria, and the animalcule Adineta steineri, not anticipated to produce SMM and/or DMSP. These diverse MmtN enzymes, alongside the larger plant MMT enzyme with an N-terminus homologous to MmtN, likely utilize the same proximity and desolvation mechanism. This study provides important insights into the catalytic mechanism of SMM and/or DMSP production, and proposes roles for these compounds in secondary metabolite production, and SMM cycling in diverse organisms and environments.

The reaction of protein-bound iron-sulfur (Fe-S) clusters with nitric oxide (NO) plays key roles in NO-mediated toxicity and signaling. Elucidation of the mechanism of the reaction of NO with DNA regulatory proteins that contain Fe-S clusters has been hampered by a lack of information about the nature of the iron-nitrosyl products formed. Herein, we report nuclear resonance vibrational spectroscopy (NRVS) and density functional theory (DFT) calculations that identify NO reaction products in WhiD and NsrR, regulatory proteins that use a [4Fe-4S] cluster to sense NO. This work reveals that nitrosylation yields multiple products structurally related to Roussin's Red Ester (RRE, [Fe2 (NO)4 (Cys)2 ]) and Roussin's Black Salt (RBS, [Fe4 (NO)7 S3 ]. In the latter case, the absence of 32 S/34 S shifts in the Fe-S region of the NRVS spectra suggest that a new species, Roussin's Black Ester (RBE), may be formed, in which one or more of the sulfide ligands is replaced by Cys thiolates.

Ferritins are multimeric cage‐forming proteins that play a crucial role in cellular iron homeostasis. All H‐chain‐type ferritins harbour a diiron site, the ferroxidase centre, at the centre of a 4 α‐helical bundle, but bacterioferritins are unique in also binding 12 hemes per 24 meric assembly. The ferroxidase centre is known to be required for the rapid oxidation of Fe2+ during deposition of an immobilised ferric mineral core within the protein's hollow interior. In contrast, the heme of bacterioferritin is required for the efficient reduction of the mineral core during iron release, but has little effect on the rate of either oxidation or mineralisation of iron. Thus, the current view is that these two cofactors function in iron uptake and release, respectively, with no functional overlap. However, rapid electron transfer between the heme and ferroxidase centre of bacterioferritin from Escherichia coli was recently demonstrated, suggesting that the two cofactors may be functionally connected. Here we report absorbance and (magnetic) circular dichroism spectroscopies, together with in vitro assays of iron‐release kinetics, which demonstrate that the ferroxidase centre plays an important role in the reductive mobilisation of the bacterioferritin mineral core, which is dependent on the heme‐ferroxidase centre electron transfer pathway. [ABSTRACT FROM AUTHOR]

The Escherichia coli fumarate and nitrate reduction (FNR) regulator acts as the cell's master switch for the transition between anaerobic and aerobic respiration, controlling the expression of >300 genes in response to O2 availability. Oxygen is perceived through a reaction with FNR's [4Fe-4S] cluster cofactor. In addition to its primary O2 signal, the FNR [4Fe-4S] cluster also reacts with nitric oxide (NO). In response to physiological concentrations of NO, FNR de-represses the transcription of hmp, which encodes a principal NO-detoxifying enzyme, and fails to activate the expression of the nitrate reductase (nar) operon, a significant source of endogenous cellular NO. Here, we show that the L28H variant of FNR, which is much less reactive towards O2 than wild-type FNR, remains highly reactive towards NO. A high resolution structure and molecular dynamics (MD) simulations of the closely related L28H-FNR from Aliivibrio fischeri revealed decreased conformational flexibility of the Cys20-Cys29 cluster-binding loop that is suggested to inhibit outer-sphere O2 reactivity, but only partially impair inner-sphere NO reactivity. Our data provide new insights into the mechanistic basis for how iron–sulfur cluster regulators can distinguish between O2 and NO. [ABSTRACT FROM AUTHOR]