The sulfo-Embden–Meyerhof–Parnas (sulfo-EMP) pathway enables Escherichia coli to utilize sulfoquinovose, (SQ) a sulfonated sugar derived from plant sulfolipids, as a carbon source. This pathway is encoded by the yih gene cassette. However, structural and functional diversity of this cassette across E. coli strains has not been fully characterized. We identified two structural variants of the yih cassette across E. coli and Shigella strains: a long form (ompL-yihOPQRSTUVW) and a truncated short form (yihTUVW). Both forms occupy the same genomic location but differ in orientation and are scattered across the phylogenetic tree, suggesting frequent recombination events. Transcriptome analyses revealed that only the long cassette, as found in E. coli K-12 MG1655, is transcriptionally induced during growth on sulfoquinovose. The short cassette, found in E. coli Nissle 1917 and other host-adapted strains, showed no differential expression. Despite this, both strains grew comparably on sulfoquinovose, indicating different metabolic adaptation strategies. Gene expression profiling revealed shared stress responses but distinct central metabolic patterns. Electrophoretic mobility shift assays further demonstrated that the transcription factor YihW from Nissle 1917 binds its DNA targets with lower affinity than the homolog from K-12 and shows weaker sulfoquinovose-dependent dissociation.

CsqR (YihW) is a local transcription factor that controls expression of yih genes involved in degradation of sulfoquinovose in Escherichia coli. We recently showed that expression of the respective gene cassette might be regulated by lactose. Here, we explore the phylogenetic and functional traits of CsqR. Phylogenetic analysis revealed that CsqR had a conserved Met25. Western blot demonstrated that CsqR was synthesized in the bacterial cell as two protein forms, 28.5 (CsqR-l) and 26 kDa (CsqR-s), the latter corresponding to start of translation at Met25. CsqR-s was dramatically activated during growth with sulfoquinovose as a sole carbon source, and displaced CsqR-l in the stationary phase during growth on rich medium. Molecular dynamic simulations revealed two possible states of the CsqR-s structure, with the interdomain linker being represented by either a disordered loop or an ɑ-helix. This helix allowed the hinge-like motion of the N-terminal domain resulting in a switch of CsqR-s between two conformational states, “open” and “compact”. We then modeled the interaction of both CsqR forms with putative effectors sulfoquinovose, sulforhamnose, sulfoquinovosyl glycerol, and lactose, and revealed that they all preferred the same pocket in CsqR-l, while in CsqR-s there were two possible options dependent on the linker structure.

Providencia is a genus of Gram-negative bacteria belonging to the Morganellaceae family. This genus includes nine species (P. stuartii, P. sneebia, P. rettgeri, P. rustigianii, P. heimbachae, P. burhodogranariea, P. alcalifaciens, P. huaxiensis , and P. vermicola ) with varying degrees of virulence, capable of infecting humans and insects [1, 2]. For Gram-negative bacteria, the somatic antigen (O-antigen) has become one of the key virulence factors. It is the highly immunogenic part of lipopolysaccharides due to the distal location. O-antigens are characterized by structural heterogeneity, providing varying degrees of inter- and intraspecific virulence. At the genetic level, somatic antigens have an operon structure. Operon genes responsible for the synthesis and transformation of O-polysaccharide are transcribed together. Analysis of O-antigen operon organization determines genes specific for each O-serogroup. It is beneficial for molecular typing of strains and for studies of bacterial evolution. This study focuses on identifying and comparing candidates for O-antigen operons in Providencia species with different levels of virulence. The hypothesis is the presence of an association between the O-antigen operon composition and the bacteria lifestyle. Data processing and analysis are carried out by a pipeline developed by the authors. Pipeline combines five steps of the genome analysis: genome quality evaluation, assembly annotation, operon identification with verification of operon boundaries, and visualization of O-antigen operons. The results reveal previously undescribed O-antigen genes and the changes in the O-antigen operons structure. Among the changes are a transposon insertion leading to tetracycline resistance and the presence of IS elements.