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      • Record: found
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      The most abundant cyst wall proteins of Acanthamoeba castellanii are lectins that bind cellulose and localize to distinct structures in developing and mature cyst walls

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          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Abstract

          Background

          Acanthamoeba castellanii, which causes keratitis and blindness in under-resourced countries, is an emerging pathogen worldwide, because of its association with contact lens use. The wall makes cysts resistant to sterilizing reagents in lens solutions and to antibiotics applied to the eye.

          Methodology/Principal findings

          Transmission electron microscopy and structured illumination microscopy (SIM) showed purified cyst walls of A. castellanii retained an outer ectocyst layer, an inner endocyst layer, and conical ostioles that connect them. Mass spectrometry showed candidate cyst wall proteins were dominated by three families of lectins (named here Jonah, Luke, and Leo), which bound well to cellulose and less well to chitin. An abundant Jonah lectin, which has one choice-of-anchor A (CAA) domain, was made early during encystation and localized to the ectocyst layer of cyst walls. An abundant Luke lectin, which has two carbohydrate-binding modules (CBM49), outlined small, flat ostioles in a single-layered primordial wall and localized to the endocyst layer and ostioles of mature walls. An abundant Leo lectin, which has two unique domains with eight Cys residues each (8-Cys), localized to the endocyst layer and ostioles. The Jonah lectin and glycopolymers, to which it binds, were accessible in the ectocyst layer. In contrast, Luke and Leo lectins and the glycopolymers, to which they bind, were mostly inaccessible in the endocyst layer and ostioles.

          Conclusions/Significance

          The most abundant A. castellanii cyst wall proteins are three sets of lectins, which have carbohydrate-binding modules that are conserved (CBM49s of Luke), newly characterized (CAA of Jonah), or unique to Acanthamoebae (8-Cys of Leo). Cyst wall formation is a tightly choreographed event, in which lectins and glycopolymers combine to form a mature wall with a protected endocyst layer. Because of its accessibility in the ectocyst layer, an abundant Jonah lectin is an excellent diagnostic target.

          Author summary

          A half century ago, investigators identified cellulose in the Acanthamoeba cyst wall, which has two layers and conical ostioles that connect them. Here we showed cyst walls contain three large sets of cellulose-binding lectins, which localize to the ectocyst layer (a Jonah lectin) or to the endocyst layer and ostioles (Luke and Leo lectins). We used the lectins to establish a sequence for cyst wall assembly when trophozoites are starved and encyst. In the first stage, a Jonah lectin and glycopolymers were present in dozens of distinct vesicles. In the second stage, a primordial wall contained small, flat ostioles outlined by a Luke lectin. In the third stage, a Jonah lectin remained in the ectocyst layer, while Luke and Leo lectins moved to the endocyst layer and ostioles. A description of the major events during cyst wall development is a starting point for mechanistic studies of its assembly.

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          Most cited references 77

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          Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.

           S Altschul (1997)
          The BLAST programs are widely used tools for searching protein and DNA databases for sequence similarities. For protein comparisons, a variety of definitional, algorithmic and statistical refinements described here permits the execution time of the BLAST programs to be decreased substantially while enhancing their sensitivity to weak similarities. A new criterion for triggering the extension of word hits, combined with a new heuristic for generating gapped alignments, yields a gapped BLAST program that runs at approximately three times the speed of the original. In addition, a method is introduced for automatically combining statistically significant alignments produced by BLAST into a position-specific score matrix, and searching the database using this matrix. The resulting Position-Specific Iterated BLAST (PSI-BLAST) program runs at approximately the same speed per iteration as gapped BLAST, but in many cases is much more sensitive to weak but biologically relevant sequence similarities. PSI-BLAST is used to uncover several new and interesting members of the BRCT superfamily.
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            Predicting transmembrane protein topology with a hidden Markov model: application to complete genomes.

            We describe and validate a new membrane protein topology prediction method, TMHMM, based on a hidden Markov model. We present a detailed analysis of TMHMM's performance, and show that it correctly predicts 97-98 % of the transmembrane helices. Additionally, TMHMM can discriminate between soluble and membrane proteins with both specificity and sensitivity better than 99 %, although the accuracy drops when signal peptides are present. This high degree of accuracy allowed us to predict reliably integral membrane proteins in a large collection of genomes. Based on these predictions, we estimate that 20-30 % of all genes in most genomes encode membrane proteins, which is in agreement with previous estimates. We further discovered that proteins with N(in)-C(in) topologies are strongly preferred in all examined organisms, except Caenorhabditis elegans, where the large number of 7TM receptors increases the counts for N(out)-C(in) topologies. We discuss the possible relevance of this finding for our understanding of membrane protein assembly mechanisms. A TMHMM prediction service is available at http://www.cbs.dtu.dk/services/TMHMM/. Copyright 2001 Academic Press.
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              • Record: found
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              • Article: found
              Is Open Access

              The carbohydrate-active enzymes database (CAZy) in 2013

              The Carbohydrate-Active Enzymes database (CAZy; http://www.cazy.org) provides online and continuously updated access to a sequence-based family classification linking the sequence to the specificity and 3D structure of the enzymes that assemble, modify and breakdown oligo- and polysaccharides. Functional and 3D structural information is added and curated on a regular basis based on the available literature. In addition to the use of the database by enzymologists seeking curated information on CAZymes, the dissemination of a stable nomenclature for these enzymes is probably a major contribution of CAZy. The past few years have seen the expansion of the CAZy classification scheme to new families, the development of subfamilies in several families and the power of CAZy for the analysis of genomes and metagenomes. This article outlines the changes that have occurred in CAZy during the past 5 years and presents our novel effort to display the resolution and the carbohydrate ligands in crystallographic complexes of CAZymes.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: InvestigationRole: MethodologyRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: InvestigationRole: MethodologyRole: Writing – original draft
                Role: ConceptualizationRole: Data curationRole: InvestigationRole: MethodologyRole: Writing – original draft
                Role: Data curationRole: InvestigationRole: Methodology
                Role: ConceptualizationRole: Funding acquisitionRole: ResourcesRole: SupervisionRole: Writing – original draft
                Role: ConceptualizationRole: Funding acquisitionRole: MethodologyRole: Project administrationRole: ResourcesRole: SupervisionRole: Writing – original draft
                Role: ConceptualizationRole: Formal analysisRole: Funding acquisitionRole: Project administrationRole: ResourcesRole: SupervisionRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS Negl Trop Dis
                PLoS Negl Trop Dis
                plos
                plosntds
                PLoS Neglected Tropical Diseases
                Public Library of Science (San Francisco, CA USA )
                1935-2727
                1935-2735
                16 May 2019
                May 2019
                : 13
                : 5
                Affiliations
                [1 ] Department of Molecular and Cell Biology, Boston University Goldman School of Dental Medicine, Boston, Massachusetts, United States of America
                [2 ] Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts, United States of America
                [3 ] Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia, United States of America
                Johns Hopkins Bloomberg School of Public Health, UNITED STATES
                Author notes

                The authors have declared that no competing interests exist.

                [¤a]

                Current address: Sarepta Therapeutics, Andover, Massachusetts, United States of America

                [¤b]

                Current address: Glyde Bio, Inc., Cambridge, Massachusetts, United States of America

                Article
                PNTD-D-19-00046
                10.1371/journal.pntd.0007352
                6541295
                31095564
                © 2019 Magistrado-Coxen et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                Counts
                Figures: 9, Tables: 1, Pages: 33
                Product
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/100000060, National Institute of Allergy and Infectious Diseases;
                Award ID: AI110638 and AI130889
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000057, National Institute of General Medical Sciences;
                Award ID: GM104603
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000015, U.S. Department of Energy;
                Award ID: DESC0015662
                Award Recipient :
                This study was funded by the National Institute of Allergy and Infectious Diseases to JS (AI110638 and AI130889), the U.S. Department of Energy to BRU (DESC0015662), and the National Institute of General Medical Sciences to CEC (GMS GM104603). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Biochemistry
                Proteins
                Lectins
                Physical Sciences
                Chemistry
                Chemical Compounds
                Organic Compounds
                Cellulose
                Physical Sciences
                Chemistry
                Organic Chemistry
                Organic Compounds
                Cellulose
                Physical Sciences
                Chemistry
                Polymer Chemistry
                Macromolecules
                Polymers
                Chitin
                Physical Sciences
                Materials Science
                Materials
                Polymers
                Chitin
                Physical Sciences
                Chemistry
                Polymer Chemistry
                Polymers
                Chitin
                Biology and Life Sciences
                Parasitology
                Parasite Groups
                Apicomplexa
                Trophozoites
                Biology and Life Sciences
                Cell Biology
                Cellular Structures and Organelles
                Vesicles
                Biology and Life Sciences
                Biochemistry
                Proteins
                Post-Translational Modification
                Signal Peptides
                Biology and Life Sciences
                Organisms
                Eukaryota
                Protozoans
                Parasitic Protozoans
                Acanthamoeba
                Biology and Life Sciences
                Organisms
                Eukaryota
                Protists
                Custom metadata
                vor-update-to-uncorrected-proof
                2019-05-29
                The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium ( http://proteomecentral.proteomexchange.org) via the PRIDE partner repository with the dataset identifier PXD011826.

                Infectious disease & Microbiology

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