Spatial mapping of immunological epitopes in the Candida cell wall using C-type lectin probes

The primary recognition event between a fungal pathogen and the immune system normally involves the engagement of a pattern recognition receptor with specific components of the cell wall. However, the cell wall is a complex three dimensional structure whose composition changes rapidly in accordance with environmental stimuli. Therefore it is important to know what is the precise nature of the primary recognition event, how many events occur to activate the immune response and how these recognition events are affected by changes in cell wall architecture, cellular morphogenesis and physiological adaptation of the pathogen to specific niches in the human body. We address this fundamental question using four soluble immune C-Type lectin receptor-probes which recognize specific mannans and β-1,3 glucan in the cell wall. We use these C-type lectin probes to demonstrate that mannan epitopes are differentially distributed in the inner and outer layers of fungal cell wall in a clustered or diffuse manner. Immune reactivity of fungal cell surfaces did not correlate with relatedness of different fungal species, and mannan-detecting receptor-probes discriminated between cell surface mannans generated by the same fungus growing under different conditions. These studies demonstrate that mannan-epitopes within fungal cell walls are differentially distributed and dynamically expressed as the fungus adapted to microenvironments that would be encountered in vivo.

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Pseudomonas Lipodepsipeptides and Fungal Cell Wall-Degrading Enzymes Act Synergistically in Biological Control

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2002.15.4.323 ◽

2002 ◽

Vol 15 (4) ◽

pp. 323-333 ◽

Cited By ~ 55

Author(s):

Vincenzo Fogliano ◽

Alessandro Ballio ◽

Monica Gallo ◽

Sheridan Woo ◽

Felice Scala ◽

...

Keyword(s):

Cell Wall ◽

Pseudomonas Syringae ◽

Enzyme Inhibitor ◽

Fungal Species ◽

Cell Wall Degrading Enzymes ◽

Fungal Cell ◽

Knock Out ◽

Degrading Enzymes ◽

Combined Action

Pseudomonas syringae pv. syringae strain B359 secreted two main lipodepsipeptides (LDPs), syringomycin E (SRE) and syringopeptin 25A (SP25A), together with at least four types of cell wall—degrading enzymes (CWDEs). In antifungal bioassays, the purified toxins SRE and SP25A interacted synergistically with chitinolytic and glucanolytic enzymes purified from the same bacterial strain or from the biocontrol fungus Trichoderma atroviride strain P1. The synergism between LDPs and CWDEs occurred against all seven different fungal species tested and P. syringae itself, with a level dependent on the enzyme used to permeabilize the microbial cell wall. The antifungal activity of SP25A was much more increased by the CWDE action than was that of the smaller SRE, suggesting a stronger antifungal role for SP25A. In vivo biocontrol assays were performed by using P. syringae alone or in combination with T. atroviride, including a Trichoderma endochitinase knock-out mutant in place of the wild type and a chitinase-specific enzyme inhibitor. These experiments clearly indicate that the synergistic interaction LDPs—CWDEs is involved in the antagonistic mechanism of P. syringae, and they support the concept that a more effective disease control is given by the combined action of the two agents.

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In vitro and in vivo polysaccharides assembly: ultrastructural and cytochemical study of growing plant cell wall components.

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100083035 ◽

1978 ◽

Vol 36 (2) ◽

pp. 434-435

Author(s):

D. Reis ◽

B. Vian ◽

J. C. Roland

Keyword(s):

Cell Wall ◽

Self Assembly ◽

Plant Cell Wall ◽

Higher Plants ◽

Three Dimensional ◽

Cytochemical Study ◽

Wall Components

Wall morphogenesis in higher plants is a problem still open to controversy. Until now the possibility of a transmembrane control and the involvement of microtubules were mostly envisaged. Self-assembly processes have been observed in the case of walls of Chlamydomonas and bacteria. Spontaneous gelling interactions between xanthan and galactomannan from Ceratonia have been analyzed very recently. The present work provides indications that some processes of spontaneous aggregation could occur in higher plants during the formation and expansion of cell wall.Observations were performed on hypocotyl of mung bean (Phaseolus aureus) for which growth characteristics and wall composition have been previously defined.In situ, the walls of actively growing cells (primary walls) show an ordered three-dimensional organization (fig. 1). The wall is typically polylamellate with multifibrillar layers alternately transverse and longitudinal. Between these layers intermediate strata exist in which the orientation of microfibrils progressively rotates. Thus a progressive change in the morphogenetic activity occurs.

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Structure of the Lectin Mannose 6-Phosphate Receptor Homology (MRH) Domain of Glucosidase II, an Enzyme That Regulates Glycoprotein Folding Quality Control in the Endoplasmic Reticulum

Journal of Biological Chemistry ◽

10.1074/jbc.m113.450239 ◽

2013 ◽

Vol 288 (23) ◽

pp. 16460-16475 ◽

Cited By ~ 21

Author(s):

Linda J. Olson ◽

Ramiro Orsi ◽

Solana G. Alculumbre ◽

Francis C. Peterson ◽

Ivan D. Stigliano ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Three Dimensional ◽

Binding Pocket ◽

Dimensional Structure ◽

Mannose Residue ◽

Glucosidase Ii ◽

Mannose 6 Phosphate ◽

First Time ◽

Conserved Residue

Here we report for the first time the three-dimensional structure of a mannose 6-phosphate receptor homology (MRH) domain present in a protein with enzymatic activity, glucosidase II (GII). GII is involved in glycoprotein folding in the endoplasmic reticulum. GII removes the two innermost glucose residues from the Glc3Man9GlcNAc2 transferred to nascent proteins and the glucose added by UDP-Glc:glycoprotein glucosyltransferase. GII is composed of a catalytic GIIα subunit and a regulatory GIIβ subunit. GIIβ participates in the endoplasmic reticulum localization of GIIα and mediates in vivo enhancement of N-glycan trimming by GII through its C-terminal MRH domain. We determined the structure of a functional GIIβ MRH domain by NMR spectroscopy. It adopts a β-barrel fold similar to that of other MRH domains, but its binding pocket is the most shallow known to date as it accommodates a single mannose residue. In addition, we identified a conserved residue outside the binding pocket (Trp-409) present in GIIβ but not in other MRHs that influences GII glucose trimming activity.

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Protein Folding: Search for Basic Physical Models

The Scientific World JOURNAL ◽

10.1100/tsw.2003.50 ◽

2003 ◽

Vol 3 ◽

pp. 623-635 ◽

Cited By ~ 3

Author(s):

Ivan Y. Torshin ◽

Robert W. Harrison

Keyword(s):

Protein Folding ◽

Tertiary Structure ◽

Three Dimensional ◽

Physical Models ◽

Dimensional Structure ◽

Computational Techniques ◽

Linear Sequence ◽

Physical Forces

How a unique three-dimensional structure is rapidly formed from the linear sequence of a polypeptide is one of the important questions in contemporary science. Apart from biological context ofin vivoprotein folding (which has been studied only for a few proteins), the roles of the fundamental physical forces in thein vitrofolding remain largely unstudied. Despite a degree of success in using descriptions based on statistical and/or thermodynamic approaches, few of the current models explicitly include more basic physical forces (such as electrostatics and Van Der Waals forces). Moreover, the present-day models rarely take into account that the protein folding is, essentially, a rapid process that produces a highly specific architecture. This review considers several physical models that may provide more direct links between sequence and tertiary structure in terms of the physical forces. In particular, elaboration of such simple models is likely to produce extremely effective computational techniques with value for modern genomics.

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Computational Protein Stabilization Can Affect Folding Energy Landscapes and Lead to Domain-Swapped Dimers

10.26434/chemrxiv.13634021.v1 ◽

2021 ◽

Author(s):

Klara Markova ◽

Antonin Kunka ◽

Klaudia Chmelova ◽

Martin Havlasek ◽

Petra Babkova ◽

...

Keyword(s):

Protein Folding ◽

Protein Design ◽

Energy Landscape ◽

Single Point ◽

Three Dimensional ◽

Protein Stabilization ◽

Dimensional Structure ◽

Evolutionary Analysis ◽

Folding Energy

The functionality of a protein depends on its unique three-dimensional structure, which is a result of the folding process when the nascent polypeptide follows a funnel-like energy landscape to reach a global energy minimum. Computer-encoded algorithms are increasingly employed to stabilize native proteins for use in research and biotechnology applications. Here, we reveal a unique example where the computational stabilization of a monomeric α/β-hydrolase enzyme (Tm = 73.5°C; ΔTm > 23°C) affected the protein folding energy landscape. Introduction of eleven single-point stabilizing mutations based on force field calculations and evolutionary analysis yielded catalytically active domain-swapped intermediates trapped in local energy minima. Crystallographic structures revealed that these stabilizing mutations target cryptic hinge regions and newly introduced secondary interfaces, where they make extensive non-covalent interactions between the intertwined misfolded protomers. The existence of domain-swapped dimers in a solution is further confirmed experimentally by data obtained from SAXS and crosslinking mass spectrometry. Unfolding experiments showed that the domain-swapped dimers can be irreversibly converted into native-like monomers, suggesting that the domain-swapping occurs exclusively in vivo. Our findings uncovered hidden protein-folding consequences of computational protein design, which need to be taken into account when applying a rational stabilization to proteins of biological and pharmaceutical interest.

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Computational Protein Stabilization Can Affect Folding Energy Landscapes and Lead to Domain-Swapped Dimers

10.26434/chemrxiv.13634021 ◽

2021 ◽

Author(s):

Klara Markova ◽

Antonin Kunka ◽

Klaudia Chmelova ◽

Martin Havlasek ◽

Petra Babkova ◽

...

Keyword(s):

Protein Folding ◽

Protein Design ◽

Energy Landscape ◽

Single Point ◽

Three Dimensional ◽

Protein Stabilization ◽

Dimensional Structure ◽

Evolutionary Analysis ◽

Folding Energy

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Cryo-Electron Tomography Reveals the Comparative Three-Dimensional Architecture of Prochlorococcus, a Globally Important Marine Cyanobacterium

Journal of Bacteriology ◽

10.1128/jb.01948-06 ◽

2007 ◽

Vol 189 (12) ◽

pp. 4485-4493 ◽

Cited By ~ 61

Author(s):

Claire S. Ting ◽

Chyongere Hsieh ◽

Sesh Sundararaman ◽

Carmen Mannella ◽

Michael Marko

Keyword(s):

Cell Wall ◽

Electron Tomography ◽

Three Dimensional ◽

Membrane System ◽

Niche Differentiation ◽

Dimensional Structure ◽

Comparative Genomic ◽

Photosynthetic Membrane ◽

Peptidoglycan Biosynthesis ◽

Cryo Electron Tomography

ABSTRACT In an age of comparative microbial genomics, knowledge of the near-native architecture of microorganisms is essential for achieving an integrative understanding of physiology and function. We characterized and compared the three-dimensional architecture of the ecologically important cyanobacterium Prochlorococcus in a near-native state using cryo-electron tomography and found that closely related strains have diverged substantially in cellular organization and structure. By visualizing native, hydrated structures within cells, we discovered that the MED4 strain, which possesses one of the smallest genomes (1.66 Mbp) of any known photosynthetic organism, has evolved a comparatively streamlined cellular architecture. This strain possesses a smaller cell volume, an attenuated cell wall, and less extensive intracytoplasmic (photosynthetic) membrane system compared to the more deeply branched MIT9313 strain. Comparative genomic analyses indicate that differences have evolved in key structural genes, including those encoding enzymes involved in cell wall peptidoglycan biosynthesis. Although both strains possess carboxysomes that are polygonal and cluster in the central cytoplasm, the carboxysomes of MED4 are smaller. A streamlined cellular structure could be advantageous to microorganisms thriving in the low-nutrient conditions characteristic of large regions of the open ocean and thus have consequences for ecological niche differentiation. Through cryo-electron tomography we visualized, for the first time, the three-dimensional structure of the extensive network of photosynthetic lamellae within Prochlorococcus and the potential pathways for intracellular and intermembrane movement of molecules. Comparative information on the near-native structure of microorganisms is an important and necessary component of exploring microbial diversity and understanding its consequences for function and ecology.

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Three-dimensional structure of the bacterial cell wall peptidoglycan

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.0510182103 ◽

2006 ◽

Vol 103 (12) ◽

pp. 4404-4409 ◽

Cited By ~ 251

Author(s):

S. O. Meroueh ◽

K. Z. Bencze ◽

D. Hesek ◽

M. Lee ◽

J. F. Fisher ◽

...

Keyword(s):

Cell Wall ◽

Bacterial Cell ◽

Three Dimensional ◽

Bacterial Cell Wall ◽

Dimensional Structure ◽

Three Dimensional Structure ◽

Cell Wall Peptidoglycan

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METHODS FOR INDICATION OF NONCULTURATED VIABLE MICROORGANISMS WHEN CONTROL OF CRITICAL POINTS OF LIVESTOCK, POULTRY AND FOOD PRODUCTION TECHNOLOGY

Problems of Veterinary Sanitation, Hygiene and Ecology ◽

10.36871/vet.san.hyg.ecol.202104010 ◽

2021 ◽

Vol 1 (4) ◽

pp. 441-447

Author(s):

Ekaterina M. Lenchenko ◽

◽

Damir I. Udavliev ◽

Inna B. Pavlova ◽

◽

...

Keyword(s):

Cell Wall ◽

Luminescence Spectrum ◽

Three Dimensional ◽

Yeast Cells ◽

Dimensional Structure ◽

Heterogeneous Structure ◽

Bacterial Cells ◽

Microscopic Fungi ◽

Gram Positive Bacteria ◽

Red Luminescence

The results of morphometric and densitometric parameters biofilms are presented, effective methods of detecting uncultivated viable microorganisms isolated from a representative sample of objects of veterinary and sanitary supervision are tested and selected. Optical, luminescent and scanning electron microscopy revealed the formation of a three-dimensional structure biofilms in the form a dense network consisting of gram-negative and gram-positive bacteria, yeast cells, hyphal and pseudohyphalic forms, surrounded by an intercellular polymer matrix. The presence hyphae of microscopic fungi causes an increase in the number of cells adhered to the substrate, microcolonies were formed from bacteria and yeast cells of microscopic fungi. The pathogenesis of the syndrome of overgrowth of microorganisms is provided by the presence of various dissociative variants, the dispersion of uncultivated bacterial cells, which gain advantages in the hyperagregation of the architectonics of heterogeneous biofilms. Multilayer membranes, vesicles, cells with a defective cell wall, spheroplasts, protoplasts, L-shapes, needle-like and giant structures, and revertant cells were identified. The dynamics of changes in the viable structures microorganisms was characterized by alternating periods of decrease and increase in the intensity of biofilm formation. When detecting the viability of microorganisms in the composition biofilms, viable and non-viable cells were differentiated – a green luminescence spectrum and a red luminescence spectrum, respectively. The dissociation of the population caused an increase in the concentration of R-dissociant cells with a higher growth rate, cell lysis was detected after 48–72 h of cultivation, a change in the ratio phenotypic forms was observed – the M-dissociant was predominant. The study of the heterogeneous structure of the population, without disturbing the natural architectonics of biofilms, revealed direct correlations (r = 0,89) between morphometric (≥90 % of the field of view) and densitometric parameters (OD). The efficiency of a nutrient medium containing pancreatic hydrolyzate, mannitol, L-asparagine and glycerol was established for the repair of the cell wall, the reversal of L-forms of microorganisms.

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Antibodies against Thrombospondin-Related Anonymous Protein Do Not Inhibit Plasmodium Sporozoite Infectivity In Vivo

Infection and Immunity ◽

10.1128/iai.68.6.3667-3673.2000 ◽

2000 ◽

Vol 68 (6) ◽

pp. 3667-3673 ◽

Cited By ~ 79

Author(s):

Soren Gantt ◽

Cathrine Persson ◽

Keith Rose ◽

Ashley J. Birkett ◽

Ruben Abagyan ◽

...

Keyword(s):

Metal Ion ◽

Competitive Inhibition ◽

Polyclonal Antibodies ◽

Three Dimensional ◽

Calcium Ionophore ◽

Plasmodium Yoelii ◽

Gliding Motility ◽

Vaccine Antigen ◽

Dimensional Structure

ABSTRACT Thrombospondin-related anonymous protein (TRAP), a candidate malaria vaccine antigen, is required for Plasmodiumsporozoite gliding motility and cell invasion. For the first time, the ability of antibodies against TRAP to inhibit sporozoite infectivity in vivo is evaluated in detail. TRAP contains an A-domain, a well-characterized adhesive motif found in integrins. We modeled here a three-dimensional structure of the TRAP A-domain of Plasmodium yoelii and located regions surrounding the MIDAS (metal ion-dependent adhesion site), the presumed business end of the domain. Mice were immunized with constructs containing these A-domain regions but were not protected from sporozoite challenge. Furthermore, monoclonal and rabbit polyclonal antibodies against the A-domain, the conserved N terminus, and the repeat region of TRAP had no effect on the gliding motility or sporozoite infectivity to mice. TRAP is located in micronemes, secretory organelles of apicomplexan parasites. Accordingly, the antibodies tested here stained cytoplasmic TRAP brightly by immunofluorescence. However, very little TRAP could be detected on the surface of sporozoites. In contrast, a dramatic relocalization of TRAP onto the parasite surface occurred when sporozoites were treated with calcium ionophore. This likely mimics the release of TRAP from micronemes when a sporozoite contacts its target cell in vivo. Contact with hepatoma cells in culture also appeared to induce the release of TRAP onto the surface of sporozoites. If large amounts of TRAP are released in close proximity to its cellular receptor(s), effective competitive inhibition by antibodies may be difficult to achieve.

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