Wzy

Aeromonas sobria strain K928 was isolated from a common carp during a Motile Aeromonas Infection/Motile Aeromonas Septicaemia disease outbreak on a Polish fish farm and classified into the new provisional PGO1 serogroup. The lipopolysaccharide of A. sobria K928 was subjected to mild acid hydrolysis, and the O-specific polysaccharide, which was isolated by gel-permeation chromatography, was studied using sugar and methylation analyses and 1 H and 13 C NMR spectroscopy. The following structure of the branched O-specific polysaccharide repeating unit of A...

PURPOSE: To investigate the mechanisms of Klebsiella pneumoniae -induced pyogenic liver abscess (PLA). METHODS: Forty-three K. pneumoniae strains from PLAs and 436 from non-PLAs were collected. Their differences were compared for virulence genes and factors, sequence types, and serotypes. Virulence genes wzi , wzy-K1 , and wzi + wzy-K1 were deleted in K. pneumoniae NTUH-K2044. Various analyses, such as transmission electron microscopy, neutrophil killing tests, and mouse lethality tests, were used to confirm the consequent changes...

The enterobacterial common antigen (ECA) is a carbohydrate polymer that is associated with the cell envelope in the Enterobacteriaceae . ECA contains a repeating trisaccharide which is polymerized by WzyE, a member of the Wzy membrane protein polymerase superfamily. WzyE activity is regulated by a membrane protein polysaccharide co-polymerase, WzzE. Förster resonance energy transfer experiments demonstrate that WzyE and WzzE from Pectobacterium atrosepticum form a complex in vivo , and immunoblotting and cryo-electron microscopy (cryo-EM) analysis confirm a defined stoichiometry of approximately eight WzzE to one WzyE...

The Pss-I region of Rhizobium leguminosarum bv. trifolii TA1 comprises more than 20 genes coding for glycosyltransferases, modifying enzymes, and polymerization/export proteins, altogether determining the biosynthesis of symbiotically relevant exopolysaccharides. In this study, the role of homologous PssG and PssI glycosyltransferases in exopolysaccharide subunit synthesis were analyzed. It was shown that the glycosyltransferase-encoding genes of the Pss-I region were part of a single large transcriptional unit with potential downstream promoters activated in specific conditions...

Nosocomial infections caused by multidrug-resistant (MDR) Enterobacter cloacae complex (ECC) pathogens are on the rise. However, the virulence strategies employed by these pathogens remain elusive. Here, we study the interaction of ECC clinical isolates with human serum to define how this pathogen evades the antimicrobial action of complement, one of the first lines of host-mediated immune defense. We identified a small number of serum-sensitive strains, including Enterobacter hormaechei strain NR3055, which we exploited for the in vitro selection of serum-resistant clones...

The O-antigen, a long polysaccharide that constitutes the distal part of the outer membrane-anchored lipopolysaccharide, is one of the critical components in the protective outer membrane of Gram-negative bacteria. Most species produce one of the structurally diverse O-antigens, with nearly all the polysaccharide components having complex structures made by the Wzx/Wzy pathway. This pathway produces repeat-units of mostly 3-8 sugars on the cytosolic face of the cytoplasmic membrane that is translocated by Wzx flippase to the periplasmic face and polymerized by Wzy polymerase to give long-chain polysaccharides...

Members of the genus Alteromonas are widely distributed in diverse marine environments and are often associated with marine organisms. Their ability to produce exopolysaccharides (EPS) and depolymerize sulfated algal polysaccharides has provided industrial importance to some species. Here, we describe the draft genome of an algae-associated strain namely, Alteromonas sp. PRIM-21 isolated from the southwest coast of India to understand the EPS biosynthetic pathways as well as polysaccharide depolymerization system in comparison to the closely related strain Alteromonas fortis 1T that shares 99...

Lactococcus lactis and Lactococcus cremoris compose commercial starter cultures widely used for industrial dairy fermentations. Some lactococcal strains may produce exopolysaccharides (EPS), which have technological applications, including texture production and phage resistance. Two distinct gene clusters associated with EPS production, designated 6073-like and 7127-like, were identified on plasmids in lactococcal strains. Infectivity of two subsets of P335 group phages, distinguished based on their single-component baseplate/receptor-binding protein nucleotide sequences, was correlated to the presence of a host-encoded 6073-like or 7127-like eps gene cluster...

Secretion of high-molecular-weight polysaccharides across the bacterial envelope is ubiquitous, as it enhances prokaryotic survival in (a)biotic settings. Such polymers are often assembled by Wzx/Wzy- or ABC transporter-dependent schemes implicating outer membrane (OM) polysaccharide export (OPX) proteins in cell-surface polymer translocation. In the social predatory bacterium Myxococcus xanthus, the exopolysaccharide (EPS) pathway WzaX, major spore coat (MASC) pathway WzaS, and biosurfactant polysaccharide (BPS) pathway WzaB were herein found to be truncated OPX homologues of Escherichia coli Wza lacking OM-spanning α-helices...

BACKGROUND: Escherichia coli O157: H7, being the cause of hemorrhagic colitis in human, is recognized as one of the most dangerous and widespread foodborne pathogens. A highly specific, sensitive and rapid E. coli O157: H7 detection method need to be developed since the traditional detection methods are complex, costly and time-consuming. OBJECTIVE: In this study, a recombinase aided amplification (RAA) assisted CRISPR/Cas12a (RAA-CRISPR/Cas12a) fluorescence platform for specific, sensitive and rapid nucleic acid detection of E...

Shigella flexneri implements the Wzy-dependent pathway to biosynthesize the O antigen (Oag) component of its surface lipopolysaccharide. The inner membrane polymerase WzySF catalyzes the repeat addition of undecaprenol-diphosphate-linked Oag (Und-PP-RUs) to produce a polysaccharide, the length of which is tightly regulated by two competing copolymerase proteins, WzzSF (short-type Oag; 10 to 17 RUs) and WzzpHS-2 (very-long-type Oag; >90 RUs). The nature of the interaction between WzySF and WzzSF /WzzpHS-2 in Oag polymerization remains poorly characterized, with the majority of the literature characterizing the individual protein constituents of the Wzy-dependent pathway...

In Gram-negative bacteria, secreted polysaccharides have multiple critical functions. In Wzx/Wzy- and ABC transporter-dependent pathways, an outer membrane (OM) polysaccharide export (OPX) type translocon exports the polysaccharide across the OM. The paradigm OPX protein Wza of Escherichia coli is an octamer in which the eight C-terminal domains form an α-helical OM pore and the eight copies of the three N-terminal domains (D1 to D3) form a periplasmic cavity. In synthase-dependent pathways, the OM translocon is a 16- to 18-stranded β-barrel protein...

Cell-free protein synthesis (CFPS) systems are emerging as powerful platforms for in vitro protein production, which leads to the development of new CFPS systems for different applications. To expand the current CFPS toolkit, here we develop a novel CFPS system derived from a chassis microorganism Klebsiella pneumoniae , an important industrial host for heterologous protein expression and the production of many useful chemicals. First, we engineered the K. pneumoniae strain by deleting a capsule formation-associated wzy gene...

The K98 capsular polysaccharide (CPS) from the Acinetobacter baumannii clinical isolate, REV-1184, was studied by sugar analysis and Smith degradation along with one- and two-dimensional 1 H and 13 C NMR spectroscopy and high-resolution electrospray ionization mass spectrometry. The CPS was found to consist of linear tetrasaccharide repeats (K-units) that include one residue each of d-GlcpNAc, d-GalpNAc, 2-acetamido-2-deoxy-d-galacturonic acid (d-GalpNAcA), and 2-acetamido-2,6-dideoxy-d-glucose (N-acetylquinovosamine, d-QuipNAc), with the GalpNAc residue decorated with a (R)-configurated 4,6-pyruvic acid acetal group...

Acinetobacter baumannii isolate LUH5552 carries the KL89 capsule biosynthesis gene cluster. Capsular polysaccharide (CPS) isolated from LUH5552 was analyzed by sugar analysis, Smith degradation, and one- and two-dimensional 1 H and 13 C NMR spectroscopy. The K89 CPS structure has not been seen before in A. baumannii CPS structures resolved to date and includes a 3-acetamido-3,6-dideoxy-d-galactose (d-Fucp3NAc) residue which is rare amongst A. baumannii CPS. The K89 CPS has a →3)-α-d-GalpNAc-(1→3)-β-d-GlcpNAc-(1→ main chain with a β-d-Glcp-(1→2)-β-d-Fucp3NAc-(1→6)-d-Glcp side branch that is α-(1→4) linked to d-GalpNAc...

Gliomas account for the majority of primary malignant brain tumors around the world and are highly aggressive. Evodiamine is one of the main effective components of Evodia rutaecarpa, which can inhibit proliferation and promote apoptosis of tumor cells including glioma cells. The derivative of Evodiamine named WZY-321 was successfully developed, and exhibited significant cytotoxicity and could efficiently induce glioma cell apoptosis; however, the mechanism of WZY-321-induced glioma cell apoptosis is not clear...

A comprehensive understanding of capsular polysaccharide (CPS) diversity is critical to implementation of phage therapy to treat panresistant Acinetobacter baumannii infections. Predictions from genome sequences can assist identification of the CPS type but can be complicated if genes outside the K locus (CPS biosynthesis gene cluster) are involved. Here, the CPS produced by A. baumannii clinical isolate 36-1454 carrying a novel K locus, KL127, was determined and compared to other CPSs. KL127 differs from KL128 in only two of the glycosyltransferase ( gtr ) genes...

Enterobacteriales have evolved a specialized outer membrane polysaccharide [Enterobacterial Common Antigen (ECA)] which allows them to persist in various environmental niches. Biosynthesis of ECA initiates on the cytoplasmic leaflet of the inner membrane (IM) where glycosyltransferases assemble ECA repeat units (RUs). Complete RUs are then translocated across the IM and assembled into polymers by ECA-specific homologues of the Wzy-dependent pathway. Consisting of the membrane proteins Wzx, Wzy and Wzz, the Wzy-dependent pathway is the most common polysaccharide biosynthetic pathway in Gram-negative bacteria where it is most notably involved in LPS O antigen (Oag) biosynthesis...

The presence of an exopolysaccharide (EPS) layer surrounding bacterial cells, termed a "glycocalyx", confers protection against toxic molecules. However, the effect of glycocalyx integrity on the tolerance to such agents is poorly understood. Using a modified disc-diffusion assay, we tested the susceptibility to a panel of antibiotics and oxidative stress-inducing compounds of various mutant strains of the social predatory Gram-negative soil bacterium Myxococcus xanthus ; the selected mutants were those that manifest different physical states of their respective EPS glycocalyces...

Shigella flexneri utilises the Wzy-dependent pathway for the production of a plethora of complex polysaccharides, including the lipopolysaccharide O-antigen (Oag) component. The inner membrane protein WzySF polymerises Oag repeat units, whilst two co-polymerase proteins, WzzSF and WzzpHS-2 , together interact with WzySF to regulate production of short- (S-Oag) and very long- (VL-Oag) Oag modal lengths, respectively. The 2D arrangement of WzySF transmembrane and soluble regions has been previously deciphered, however, attaining information on the 3D structural and conformational arrangement of WzySF, or any homologue, has proven difficult...