Heterologous expression of a set of genes present on a subgenomic fragment of S. davawensis resulted in the production of roseoflavin by the host Streptomyces coelicolor M1152. Phylogenetic analysis revealed that S. davawensis is a close relative of Streptomyces cinnabarinus, and much to our surprise, we found that the latter bacterium is a roseoflavin producer as well.

The bacteria Streptomyces davawensis and Streptomyces cinnabarinus produce roseoflavin, the only known natural riboflavin (vitamin B2 ) analogue with antibiotic activity. Roseoflavin can be considered a natural antimetabolite and has been postulated to be biosynthesized from riboflavin via the key intermediate 8-demethyl-8-aminoriboflavin (AF). The required site-specific substitution of one of the methyl groups on the dimethylbenzene ring of riboflavin by an amino group (to give AF) is challenging. The pathway from riboflavin to AF has remained elusive, and the corresponding enzyme/s was/were unknown. Herein, we show by systematic gene deletion, heterologous gene expression, and biochemical studies that the enzyme specified by the gene BN159_7989 from S. davawensis is able to carry out a whole set of chemical reactions starting from riboflavin-5'-phosphate to give the final product 8-demethyl-8-aminoriboflavin-5'-phosphate (AFP).

FMN riboswitches are genetic elements that, in many bacteria, control genes responsible for biosynthesis and/or transport of riboflavin (vitamin B 2 ). We report that the Escherichia coli ribB FMN riboswitch controls expression of the essential gene ribB coding for the riboflavin biosynthetic enzyme 3,4-dihydroxy-2-butanone-4-phosphate synthase (RibB; EC 4.1.99.12). Our data show that the E. coli ribB FMN riboswitch is unusual because it operates at the transcriptional and also at the translational level. Expression of ribB is negatively affected by FMN and by the FMN analog roseoflavin mononucleotide, which is synthesized enzymatically from roseoflavin and ATP. Consequently, in addition to flavoenzymes, the E. coli ribB FMN riboswitch constitutes a target for the antibiotic roseoflavin produced by Streptomyces davawensis.

HbpS is an extracellular oligomeric protein, which has been shown to act in concert with the two-component system SenSSenR during the sensing of redox stress. HbpS can bind and degrade heme under oxidative stress conditions, leading to a free iron ion. The liberated iron is subsequently coordinated on the protein surface. Furthermore, HbpS has been shown to modulate the phosphorylation state of the sensor kinase SenS as, in the absence of oxidative stress conditions, HbpS inhibits SenS autophosphorylation whereas the presence of heme or iron ions and redox-stressing agents enhances it. Using HbpS wild type and mutants as well as different biochemical and biophysical approaches, we show that iron-mediated oxidative stress induces both secondary structure and overall intrinsic conformational changes within HbpS. We demonstrate in addition that HbpS is oxidatively modified, leading to the generation of highly reactive carbonyl groups and tyrosine-tyrosine bonds. Further examination of the crystal structure and subsequent mutational analyses allowed the identification of the tyrosine residue participating in dityrosine formation, which occurs between two monomers within the octomeric assembly. Therefore, it is proposed that oxidative modifications causing structural and conformational changes are responsible for the control of SenS and hence of the HbpS-SenS-SenR signaling cascade.Iron is the fourth most abundant element in the Earth's crust and is an essential trace mineral for nearly all known organisms. Under physiological conditions, it exists either in the reduced Fe 2ϩ (ferrous) or in the oxidized Fe 3ϩ (ferric) form. It plays a crucial role in many biological processes, as photosynthesis, N 2 fixation, H 2 production and consumption, respiration, oxygen transport, or gene regulation. Because of its redox potential ranging from Ϫ300 to ϩ700 mV, iron is a versatile prosthetic component that can be incorporated into proteins either as a mono-or binuclear species, or in a more complex form as part of iron-sulfur clusters or heme groups (1-3).In the presence of oxygen, iron ions frequently lead to the formation of redox stress by the generation of reactive oxygen species ( ROS can provoke the damage of DNA, lipids, and proteins (4 -6). For instance, when proteins are exposed to ROS, they undergo a variety of oxidative modifications including: Nitration of aromatic amino acid residues, hydroxylation of aromatic groups, and aliphatic amino acid side-chains, sulfoxidation of methionine residues, and conversion of some amino acid residues to carbonyl derivatives. Oxidation can also induce the cleavage of the polypeptide chain and the formation of crosslinked protein derivatives (7,8). These modifications can lead to functional changes of proteins that subsequently disturb the cellular metabolism. Thus, while bacteria and other organisms have to ensure that enough iron ions are present for the diverse biochemical reactions, they also have to avoid their harmful effects.We have previously identified the two-component sy...

Riboflavin analogs have a good potential to serve as basic structures for the development of novel anti-infectives. Riboflavin analogs have multiple cellular targets, since riboflavin (as a precursor to flavin cofactors) is active at more than one site in the cell. As a result, the frequency of developing resistance to antimicrobials based on riboflavin analogs is expected to be significantly lower. The only known natural riboflavin analog with antibiotic function is roseoflavin from the bacterium Streptomyces davawensis. This antibiotic negatively affects flavoenzymes and FMN riboswitches. Another roseoflavin producer, Streptomyces cinnabarinus, was recently identified. Possibly, flavin analogs with antibiotic activity are more widespread than anticipated. The same could be true for flavin analogs yet to be discovered, which could constitute tools for cellular chemistry, thus allowing a further extension of the catalytic spectrum of flavoenzymes.