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A structural and biochemical comparison of Ribonuclease E homologues from pathogenic bacteria highlights species-specific properties.

ABSTRACT: Regulation of gene expression through processing and turnover of RNA is a key mechanism that allows bacteria to rapidly adapt to changing environmental conditions. Consequently, RNA degrading enzymes (ribonucleases; RNases) such as the endoribonuclease RNase E, frequently play critical roles in pathogenic bacterial virulence and are potential antibacterial targets. RNase E consists of a highly conserved catalytic domain and a variable non-catalytic domain that functions as the structural scaffold for the multienzyme degradosome complex. Despite conservation of the catalytic domain, a recent study identified differences in the response of RNase E homologues from different species to the same inhibitory compound(s). While RNase E from Escherichia coli has been well-characterised, far less is known about RNase E homologues from other bacterial species. In this study, we structurally and biochemically characterise the RNase E catalytic domains from four pathogenic bacteria: Yersinia pestis, Francisella tularensis, Burkholderia pseudomallei and Acinetobacter baumannii, with a view to exploiting RNase E as an antibacterial target. Bioinformatics, small-angle x-ray scattering and biochemical RNA cleavage assays reveal globally similar structural and catalytic properties. Surprisingly, subtle species-specific differences in both structure and substrate specificity were also identified that may be important for the development of effective antibacterial drugs targeting RNase E.

SUBMITTER: Mardle CE

PROVIDER: S-EPMC6538622 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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A structural and biochemical comparison of Ribonuclease E homologues from pathogenic bacteria highlights species-specific properties.

Mardle Charlotte E CE Shakespeare Thomas J TJ Butt Louise E LE Goddard Layla R LR Gowers Darren M DM Atkins Helen S HS Vincent Helen A HA Callaghan Anastasia J AJ

Scientific reports 20190528 1

Regulation of gene expression through processing and turnover of RNA is a key mechanism that allows bacteria to rapidly adapt to changing environmental conditions. Consequently, RNA degrading enzymes (ribonucleases; RNases) such as the endoribonuclease RNase E, frequently play critical roles in pathogenic bacterial virulence and are potential antibacterial targets. RNase E consists of a highly conserved catalytic domain and a variable non-catalytic domain that functions as the structural scaffol ...[more]

PMID: 31138855

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Project description:BackgroundPeptidase family A1, to which pepsin belongs, had been assumed to be restricted to eukaryotes. The tertiary structure of pepsin shows two lobes with similar folds and it has been suggested that the gene has arisen from an ancient duplication and fusion event. The only sequence similarity between the lobes is restricted to the motif around the active site aspartate and a hydrophobic-hydrophobic-Gly motif. Together, these contribute to an essential structural feature known as a psi-loop. There is one such psi-loop in each lobe, and so each lobe presents an active Asp. The human immunodeficiency virus peptidase, retropepsin, from peptidase family A2 also has a similar fold but consists of one lobe only and has to dimerize to be active. All known members of family A1 show the bilobed structure, but it is unclear if the ancestor of family A1 was similar to an A2 peptidase, or if the ancestral retropepsin was derived from a half-pepsin gene. The presence of a pepsin homologue in a prokaryote might give insights into the evolution of the pepsin family.ResultsHomologues of the aspartic peptidase pepsin have been found in the completed genomic sequences from seven species of bacteria. The bacterial homologues, unlike those from eukaryotes, do not possess signal peptides, and would therefore be intracellular acting at neutral pH. The bacterial homologues have Thr218 replaced by Asp, a change which in renin has been shown to confer activity at neutral pH. No pepsin homologues could be detected in any archaean genome.ConclusionThe peptidase family A1 is found in some species of bacteria as well as eukaryotes. The bacterial homologues fall into two groups, one from oceanic bacteria and one from plant symbionts. The bacterial homologues are all predicted to be intracellular proteins, unlike the eukaryotic enzymes. The bacterial homologues are bilobed like pepsin, implying that if no horizontal gene transfer has occurred the duplication and fusion event might be very ancient indeed, preceding the divergence of bacteria and eukaryotes. It is unclear whether all the bacterial homologues are derived from horizontal gene transfer, but those from the plant symbionts probably are. The homologues from oceanic bacteria are most closely related to memapsins (or BACE-1 and BACE-2), but are so divergent that they are close to the root of the phylogenetic tree and to the division of the A1 family into two subfamilies.

Project description:The plant cuticle, which consists of cutin and waxes, forms a hydrophobic coating covering the aerial surfaces of all plants. It acts as an interface between plants and their surrounding environment whilst also protecting them against biotic and abiotic stresses. In this research, we have investigated the biodiversity and cuticle properties of aquatic plant duckweed, using samples isolated from four different locations around Hongze lake in Jiangsu province, China. The samples were genotyped using two chloroplast markers and nuclear ribosomal DNA markers, which revealed them as ecotypes of the larger duckweed, Spirodela polyrhiza. Duckweed cuticle properties were investigated by compositional analysis using Gas Chromatography coupled with Mass Spectroscopy (GC-MS) Flame Ionization Detector (GC-FID), and ultrastructural observation by cryo-Scanning Electron Microscopy (cryo-SEM). Cuticle compositional analysis indicated that fatty acids and primary alcohols, the two typical constituents found in many land plant cuticle, are the major duckweed wax components. A large portion of the duckweed wax fraction is composed of phytosterols, represented by campesterol, stigmasterol, sitosterol and their common precursor squalene. The cryo-SEM observation uncovered significant differences between the surface structures of the top air-facing and bottom water-facing sides of the plant fronds. The top side of the fronds, containing multiple stomata complexes, appeared to be represented by a rather flat waxy film sporadically covered with wax crystals. Underneath the waxy film was detected a barely distinguished nanoridge net, which became distinctly noticeable after chloroform treatment. On the bottom side of the fronds, the large epidermal cells were covered by the well-structured net, whose sections became narrower and sharper under cryo-SEM following chloroform treatment. These structural differences between the abaxial and adaxial sides of the fronds evidently relate to their distinct physiological roles in interacting with the contrasting environments of sunlight/air and nutrients/water. The unique structural and biochemical features of Spirodela frond surfaces with their rapid reproductive cycle and readily availability genome sequence, make duckweed an attractive monocot model for studying the fundamental processes related to plant protection against ultraviolet irradiation, pathogens and other environmental stresses.

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A structural and biochemical comparison of Ribonuclease E homologues from pathogenic bacteria highlights species-specific properties.

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A structural and biochemical comparison of Ribonuclease E homologues from pathogenic bacteria highlights species-specific properties.

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