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      Lipopolysaccharide-Linked Enterobacterial Common Antigen (ECA LPS) Occurs in Rough Strains of Escherichia coli R1, R2, and R4

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          Abstract

          Enterobacterial common antigen (ECA) is a conserved surface antigen characteristic for Enterobacteriaceae. It is consisting of trisaccharide repeating unit, →3)-α- d-Fuc p4NAc-(1→4)-β- d-Man pNAcA-(1→4)-α- d-Glc pNAc-(1→, where prevailing forms include ECA linked to phosphatidylglycerol (ECA PG) and cyclic ECA (ECA CYC). Lipopolysaccharide (LPS)-associated form (ECA LPS) has been proved to date only for rough Shigella sonnei phase II. Depending on the structure organization, ECA constitutes surface antigen (ECA PG and ECA LPS) or maintains the outer membrane permeability barrier (ECA CYC). The existence of LPS was hypothesized in the 1960–80s on the basis of serological observations. Only a few Escherichia coli strains (i.e., R1, R2, R3, R4, and K-12) have led to the generation of anti-ECA antibodies upon immunization, excluding ECA PG as an immunogen and conjecturing ECA LPS as the only immunogenic form. Here, we presented a structural survey of ECA LPS in E. coli R1, R2, R3, and R4 to correlate previous serological observations with the presence of ECA LPS. The low yields of ECA LPS were identified in the R1, R2, and R4 strains, where ECA occupied outer core residues of LPS that used to be substituted by O-specific polysaccharide in the case of smooth LPS. Previously published observations and hypotheses regarding the immunogenicity and biosynthesis of ECA LPS were discussed and correlated with presented herein structural data.

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          NMRFAM-SPARKY: enhanced software for biomolecular NMR spectroscopy

          Summary: SPARKY (Goddard and Kneller, SPARKY 3) remains the most popular software program for NMR data analysis, despite the fact that development of the package by its originators ceased in 2001. We have taken over the development of this package and describe NMRFAM-SPARKY, which implements new functions reflecting advances in the biomolecular NMR field. NMRFAM-SPARKY has been repackaged with current versions of Python and Tcl/Tk, which support new tools for NMR peak simulation and graphical assignment determination. These tools, along with chemical shift predictions from the PACSY database, greatly accelerate protein side chain assignments. NMRFAM-SPARKY supports automated data format interconversion for interfacing with a variety of web servers including, PECAN , PINE, TALOS-N, CS-Rosetta, SHIFTX2 and PONDEROSA-C/S. Availability and implementation: The software package, along with binary and source codes, if desired, can be downloaded freely from http://pine.nmrfam.wisc.edu/download_packages.html. Instruction manuals and video tutorials can be found at http://www.nmrfam.wisc.edu/nmrfam-sparky-distribution.htm. Contact: whlee@nmrfam.wisc.edu or markley@nmrfam.wisc.edu Supplementary information: Supplementary data are available at Bioinformatics online.
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            GlycoWorkbench: a tool for the computer-assisted annotation of mass spectra of glycans.

            Mass spectrometry is the main analytical technique currently used to address the challenges of glycomics as it offers unrivalled levels of sensitivity and the ability to handle complex mixtures of different glycan variations. Determination of glycan structures from analysis of MS data is a major bottleneck in high-throughput glycomics projects, and robust solutions to this problem are of critical importance. However, all the approaches currently available have inherent restrictions to the type of glycans they can identify, and none of them have proved to be a definitive tool for glycomics. GlycoWorkbench is a software tool developed by the EUROCarbDB initiative to assist the manual interpretation of MS data. The main task of GlycoWorkbench is to evaluate a set of structures proposed by the user by matching the corresponding theoretical list of fragment masses against the list of peaks derived from the spectrum. The tool provides an easy to use graphical interface, a comprehensive and increasing set of structural constituents, an exhaustive collection of fragmentation types, and a broad list of annotation options. The aim of GlycoWorkbench is to offer complete support for the routine interpretation of MS data. The software is available for download from: http://www.eurocarbdb.org/applications/ms-tools.
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              Synthesis of bacterial polysaccharides via the Wzx/Wzy-dependent pathway.

              The surfaces of bacteria mediate a multitude of functions in the environment and in an infected host, including adhesion to both biotic and abiotic substrata, motility, immune system interaction and (or) activation, biofilm formation, and cell-cell communication, with many of these features directly influenced by cell-surface glycans. In both Gram-negative and Gram-positive bacteria, the majority of cell-surface polysaccharides are produced via the Wzx/Wzy-dependent assembly pathway; these glycans include heteropolymeric O-antigen, enterobacterial common antigen, exopolysaccharide, spore coat, and capsule in diverse bacteria. The key components of this assembly pathway are the integral inner membrane Wzx flippase, Wzy polymerase, and Wzz chain-length regulator proteins, which until recently have resisted detailed structural and functional characterization. In this review, we have provided a comprehensive synthesis of the latest structural and mechanistic data for each protein, as well as an examination of substrate specificity for each assembly step and complex formation between the constituent proteins. To complement the unprecedented explosion of genomic-sequencing data for bacteria, we have also highlighted both classical and state-of-the-art methods by which encoded Wzx, Wzy, and Wzz proteins can be reliably identified and annotated, using the model Gram-negative bacterium Pseudomonas aeruginosa as an example data set. Lastly, we outline future avenues of research, with the aim of stimulating researchers to take the next steps in investigating the function of, and interplay between, the constituents of this widespread assembly scheme.
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                Author and article information

                Journal
                Int J Mol Sci
                Int J Mol Sci
                ijms
                International Journal of Molecular Sciences
                MDPI
                1422-0067
                21 August 2020
                September 2020
                : 21
                : 17
                : 6038
                Affiliations
                Laboratory of Microbial Immunochemistry and Vaccines, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Weigla 12, 53-114 Wroclaw, Poland; anna.maciejewska@ 123456hirszfeld.pl (A.M.); marta.kaszowska@ 123456hirszfeld.pl (M.K.); wojciech.jachymek@ 123456hirszfeld.pl (W.J.); czeslaw.lugowski@ 123456hirszfeld.pl (C.L.)
                Author notes
                Author information
                https://orcid.org/0000-0003-4057-2997
                https://orcid.org/0000-0001-8963-3020
                https://orcid.org/0000-0001-8081-7261
                Article
                ijms-21-06038
                10.3390/ijms21176038
                7504096
                32839412
                70a4da43-f5d2-4c2d-849a-83b0b5bbd1a6
                © 2020 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 24 July 2020
                : 20 August 2020
                Categories
                Article

                Molecular biology
                enterobacterial common antigen,eca,lipopolysaccharide,lps,ecalps,endotoxin,nmr,mass spectrometry

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