Correction to: In silico structural homology modelling of EST073 motif coding protein of tea Camellia sinensis (L)

An amendment to this paper has been published and can be accessed via the original article.

Unmasking of Gitelman Syndrome during Pregnancy in an Adolescent with Thyrotoxic Crisis

Background. Gitelman syndrome (GS) is an inherited salt-losing renal tubulopathy characterized by hypokalemic metabolic alkalosis with hypomagnesemia and hypocalciuria. Patients can be asymptomatic until late adolescence or adulthood, and hence may be discovered incidentally during presentation with other illnesses. GS has been described in association with thyroid disorders and should be considered in patients with hyperthyroidism and persistent hypokalemia, especially in those with associated hypomagnesemia and hypocalciuria. Case summary. In this report, we describe an 18-year-old female who presented with hyperemesis gravidarum and thyrotoxicosis, and was incidentally found to have GS, confirmed by the sequence analysis of SLC12A3. Conclusions. Thyroid dysfunctions, such as hypothyroidism, thyrotoxicosis, and thyroid nodules, may develop during pregnancy. A structural homology between the beta-human chorionic gonadotropin and thyroid stimulating hormone molecules, as well as their receptors is probably the basis for the transient thyrotoxicosis crisis during pregnancy. Since hyperemesis in pregnancy can also lead to hypokalemia and alkalosis, a high index of suspicion for GS during pregnancy is required for timely diagnosis and management.

The Role of Decorin and Biglycan Signaling in Tumorigenesis

The complex and adaptive nature of malignant neoplasm constitute a major challenge for the development of effective anti-oncogenic therapies. Emerging evidence has uncovered the pivotal functions exerted by the small leucine-rich proteoglycans, decorin and biglycan, in affecting tumor growth and progression. In their soluble forms, decorin and biglycan act as powerful signaling molecules. By receptor-mediated signal transduction, both proteoglycans modulate key processes vital for tumor initiation and progression, such as autophagy, inflammation, cell-cycle, apoptosis, and angiogenesis. Despite of their structural homology, these two proteoglycans interact with distinct cell surface receptors and thus modulate distinct signaling pathways that ultimately affect cancer development. In this review, we summarize growing evidence for the complex roles of decorin and biglycan signaling in tumor biology and address potential novel therapeutic implications.

Kite-Shaped Molecules Block SARS-CoV-2 Cell Entry at a Post-Attachment Step

Anti-viral small molecules are currently lacking for treating coronavirus infection. The long development timescales for such drugs are a major problem, but could be shortened by repurposing existing drugs. We therefore screened a small library of FDA-approved compounds for potential severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) antivirals using a pseudovirus system that allows a sensitive read-out of infectivity. A group of structurally-related compounds, showing moderate inhibitory activity with IC50 values in the 2–5 μM range, were identified. Further studies demonstrated that these “kite-shaped” molecules were surprisingly specific for SARS-CoV-1 and SARS-CoV-2 and that they acted early in the entry steps of the viral infectious cycle, but did not affect virus attachment to the cells. Moreover, the compounds were able to prevent infection in both kidney- and lung-derived human cell lines. The structural homology of the hits allowed the production of a well-defined pharmacophore that was found to be highly accurate in predicting the anti-viral activity of the compounds in the screen. We discuss the prospects of repurposing these existing drugs for treating current and future coronavirus outbreaks.

The mechanisms of cell invasion in cnidarian-dinoflagellate symbiosis: Learning from parasitic strategies

<p>Cnidarians such as corals, anemones and hydroids commonly form an intracellular symbiosis with photosynthetic dinoflagellates of the genus Symbiodinium. Dinoflagellate symbionts are most often obtained anew from the environment during larval development, and, once acquired reside inside host-derived vacuoles within the cnidarian gastrodermal cells. In order to gain entry to host cells, the symbionts likely interact with innate immune receptors in the extracellular matrix, the first line of defense against microbial attack. While several innate immune pathways have been described in cnidarians, little is known about the specific receptor-ligand interactions that allow the symbiont to gain entry to host cells. Furthermore, it is unclear how these pathways are involved in enabling friendly microbes to reside within host cells while maintaining an immune response to harmful pathogens. The invasion strategies of vertebrate intracellular parasites are well studied, especially those used by members of the Apicomplexa. Apicomplexan parasites have evolved mechanisms to evade immune receptors in the extracellular matrix and exploit specific receptors to their own benefit, to gain entry to host cells. Apicomplexans are closely related to dinoflagellates, both belonging to the infrakingdom Alveolata. The malaria parasite Plasmodium spp. has evolved the thromobospondin-related anonymous protein, or TRAP, that uses a thrombospondin structural homology repeat (TSR) domain to bind to a scavenger receptor (SRB1) on the hepatocyte cell surface and gain entry to the cell. This is of particular interest, as class B scavenger receptors are upregulated in the symbiotic state of two anemone species. The aims of the research presented in this thesis were to: (1) characterize the scavenger receptor (SR) repertoire in cnidarians; (2) characterize the TSR-domain-containing protein repertoire of cnidarians and their symbiotic dinoflagellates; and (3) establish, through experimental manipulation, the potential role for SR-TSR domain interactions at the onset of symbiosis in the sea anemone Aiptasia sp, a model system for the study of the cnidariandinoflagellate symbiosis. In Chapter 2, I characterized the large and diverse SR repertoire of six cnidarian species. Cnidarians lack the classic SR type-A collagen domain-containing proteins that are common in humans, however the cnidarian SR cysteine-rich domain-containing protein repertoire is expanded and diverse. Phylogenetic analysis of SR type-B proteins defines two or three distinct groups. Functional experimental data presented here show that blocking SR binding sites with fucoidan significantly reduces dinoflagellate uptake by the anemone Aiptasia sp. These data provide further evidence that SRs are important to symbiont recognition and uptake, and may be an essential component of symbiont acquisition. In Chapter 3, I investigated a SR ligand, the thrombospondin structural homology repeat, or TSR domain. In particular, I characterized the TSR-domain-containing protein repertoire of six cnidarian species and compared these proteins to vertebrate TSR proteins of known function. Searches revealed a large repertoire of TSR-domaincontaining proteins. Of particular interest is the large number of Adams metalloprotease-like proteins, a group that is common in both humans and cnidarians, suggesting that this is an ancestral TSR protein group. Phylogenetic analysis of TSR domains shows that binding motifs and 3-D folding sites are highly conserved. These data suggest that TSR domains are ancient and have changed very little in amino acid sequence from lower metazoans to vertebrates. In Chapter 4, I explored the role of TSR-domain-containing proteins at the onset of symbiosis in the model Aiptasia sp. system. In functional experiments, aposymbiotic anemones were challenged with proteins and antibodies to either block or stimulate TSR domain binding. Symbiont uptake was measured over several time-points to determine the effects on symbiont acquisition. Adding an excess of TSR-domaincontaining protein or TSR synthetic peptide increased symbiont uptake, while blocking TSR domains prevented symbiont uptake. Finally, the addition of exogenous TGFβ to TSR antibody-challenged anemones, reversed the blocking effect. These data suggest that the immune-suppressive TGFβ pathway is involved in early onset of the symbiosis. Since the TSR domain is implicated in the TGFβ pathway, these results support previous findings of the involvement of TGFβ in promoting tolerance of symbionts within the host. Apicomplexan parasites exploit scavenger receptor-TSR domain-binding to gain entry, and also use immune modulation to persist inside host cells. Data presented here suggest that dinoflagellates are utilizing the same mechanisms to form a mutualistic relationship with the cnidarian host. Overall, the work presented here provides new information about several cnidarian extracellular matrix proteins, with searches revealing large repertoires of both scavenger receptors and TSR-domain-containing proteins. Functional data suggest that both protein families are involved in the cnidarian-dinoflagellate symbiosis. Searches of the dinoflagellate genome did not find a clear dinoflagellate homologue to the apicomplexan TRAP proteins. However, this research provides further evidence that similar receptor-ligand interactions are involved in the entry of both beneficial and pathogenic microbes to host cells. These results add to growing knowledge about the complex molecular pathways that enable and support cnidarian-dinoflagellate symbiosis. An understanding of the mechanisms that support healthy symbiosis is essential when trying to predict the vitality and productivity of reef ecosystems in the face of climate change.</p>

The mechanisms of cell invasion in cnidarian-dinoflagellate symbiosis: Learning from parasitic strategies

<p>Cnidarians such as corals, anemones and hydroids commonly form an intracellular symbiosis with photosynthetic dinoflagellates of the genus Symbiodinium. Dinoflagellate symbionts are most often obtained anew from the environment during larval development, and, once acquired reside inside host-derived vacuoles within the cnidarian gastrodermal cells. In order to gain entry to host cells, the symbionts likely interact with innate immune receptors in the extracellular matrix, the first line of defense against microbial attack. While several innate immune pathways have been described in cnidarians, little is known about the specific receptor-ligand interactions that allow the symbiont to gain entry to host cells. Furthermore, it is unclear how these pathways are involved in enabling friendly microbes to reside within host cells while maintaining an immune response to harmful pathogens. The invasion strategies of vertebrate intracellular parasites are well studied, especially those used by members of the Apicomplexa. Apicomplexan parasites have evolved mechanisms to evade immune receptors in the extracellular matrix and exploit specific receptors to their own benefit, to gain entry to host cells. Apicomplexans are closely related to dinoflagellates, both belonging to the infrakingdom Alveolata. The malaria parasite Plasmodium spp. has evolved the thromobospondin-related anonymous protein, or TRAP, that uses a thrombospondin structural homology repeat (TSR) domain to bind to a scavenger receptor (SRB1) on the hepatocyte cell surface and gain entry to the cell. This is of particular interest, as class B scavenger receptors are upregulated in the symbiotic state of two anemone species. The aims of the research presented in this thesis were to: (1) characterize the scavenger receptor (SR) repertoire in cnidarians; (2) characterize the TSR-domain-containing protein repertoire of cnidarians and their symbiotic dinoflagellates; and (3) establish, through experimental manipulation, the potential role for SR-TSR domain interactions at the onset of symbiosis in the sea anemone Aiptasia sp, a model system for the study of the cnidariandinoflagellate symbiosis. In Chapter 2, I characterized the large and diverse SR repertoire of six cnidarian species. Cnidarians lack the classic SR type-A collagen domain-containing proteins that are common in humans, however the cnidarian SR cysteine-rich domain-containing protein repertoire is expanded and diverse. Phylogenetic analysis of SR type-B proteins defines two or three distinct groups. Functional experimental data presented here show that blocking SR binding sites with fucoidan significantly reduces dinoflagellate uptake by the anemone Aiptasia sp. These data provide further evidence that SRs are important to symbiont recognition and uptake, and may be an essential component of symbiont acquisition. In Chapter 3, I investigated a SR ligand, the thrombospondin structural homology repeat, or TSR domain. In particular, I characterized the TSR-domain-containing protein repertoire of six cnidarian species and compared these proteins to vertebrate TSR proteins of known function. Searches revealed a large repertoire of TSR-domaincontaining proteins. Of particular interest is the large number of Adams metalloprotease-like proteins, a group that is common in both humans and cnidarians, suggesting that this is an ancestral TSR protein group. Phylogenetic analysis of TSR domains shows that binding motifs and 3-D folding sites are highly conserved. These data suggest that TSR domains are ancient and have changed very little in amino acid sequence from lower metazoans to vertebrates. In Chapter 4, I explored the role of TSR-domain-containing proteins at the onset of symbiosis in the model Aiptasia sp. system. In functional experiments, aposymbiotic anemones were challenged with proteins and antibodies to either block or stimulate TSR domain binding. Symbiont uptake was measured over several time-points to determine the effects on symbiont acquisition. Adding an excess of TSR-domaincontaining protein or TSR synthetic peptide increased symbiont uptake, while blocking TSR domains prevented symbiont uptake. Finally, the addition of exogenous TGFβ to TSR antibody-challenged anemones, reversed the blocking effect. These data suggest that the immune-suppressive TGFβ pathway is involved in early onset of the symbiosis. Since the TSR domain is implicated in the TGFβ pathway, these results support previous findings of the involvement of TGFβ in promoting tolerance of symbionts within the host. Apicomplexan parasites exploit scavenger receptor-TSR domain-binding to gain entry, and also use immune modulation to persist inside host cells. Data presented here suggest that dinoflagellates are utilizing the same mechanisms to form a mutualistic relationship with the cnidarian host. Overall, the work presented here provides new information about several cnidarian extracellular matrix proteins, with searches revealing large repertoires of both scavenger receptors and TSR-domain-containing proteins. Functional data suggest that both protein families are involved in the cnidarian-dinoflagellate symbiosis. Searches of the dinoflagellate genome did not find a clear dinoflagellate homologue to the apicomplexan TRAP proteins. However, this research provides further evidence that similar receptor-ligand interactions are involved in the entry of both beneficial and pathogenic microbes to host cells. These results add to growing knowledge about the complex molecular pathways that enable and support cnidarian-dinoflagellate symbiosis. An understanding of the mechanisms that support healthy symbiosis is essential when trying to predict the vitality and productivity of reef ecosystems in the face of climate change.</p>

Inhibitors of Discoidin Domain Receptor (DDR) Kinases for Cancer and Inflammation

The discoidin domain receptor tyrosine kinases DDR1 and DDR2 are distinguished from other kinase enzymes by their extracellular domains, which interact with collagen rather than with peptidic growth factors, before initiating signaling via tyrosine phosphorylation. They share significant sequence and structural homology with both the c-Kit and Bcr-Abl kinases, and so many inhibitors of those kinases are also effective. Nevertheless, there has been an extensive research effort to develop potent and specific DDR inhibitors. A key interaction for many of these compounds is H-bonding to Met-704 in a hydrophobic pocket of the DDR enzyme. The most widespread use of DDR inhibitors has been for cancer therapy, but they have also shown effectiveness in animal models of inflammatory conditions such as Alzheimer’s and Parkinson’s diseases, and in chronic renal failure and glomerulonephritis.

Reshaping the binding channel of a novel GH113 family β-Mannanase from Paenibacillus cineris (PcMan113) for enhanced activity

Endo-β-mannanases are an important enzyme for degrading lignocellulosic biomass to generate mannan, which has significant health effects as a prebiotic that promotes the development of gut microbiota. Here, a novel endo-β-mannanase belonging to glycoside hydrolase (GH) family 113 from Paenibacillus cineris (PcMan113) was cloned, expressed and characterized, as one of only a few reported GH113 family β-mannanases. Compared to other functionally and structurally characterized GH113 mannanases, recombinant PcMan113 showed a broader substrate spectrum and a better performance. Based on a structural homology model, the highly active mutant PcMT3 (F110E/N246Y) was obtained, with 4.60- and 5.53-fold increases of enzyme activity (towards KG) and catalytic efficiency (kcat/Km, against M5) compared with the WT enzyme, respectively. Furthermore, molecular dynamics (MD) simulations were conducted to precisely explore the differences of catalytic activity between WT and PcMT3, which revealed that PcMT3 has a less flexible conformation, as well as an enlarged substrate binding channel with decreased steric hindrance and increased binding energy in substrate recognition. In conclusion, we obtained a highly active variant of PcMan113 with potential for commercial application in the manufacture of mannooligosaccharides.

Importance of Cry Proteins in Biotechnology: Initially a Bioinsecticide, Now a Vaccine Adjuvant

A hallmark of Bacillus thuringiensis bacteria is the formation of one or more parasporal crystal (Cry) proteins during sporulation. The toxicity of these proteins is highly specific to insect larvae, exerting lethal effects in different insect species but not in humans or other mammals. The aim of this review is to summarize previous findings on Bacillus thuringiensis, including the characteristics of the bacterium, its subsequent contribution to biotechnology as a bioinsecticide due to the presence of Cry proteins, and its potential application as an adjuvant. In several studies, Cry proteins have been administered together with specific antigens to immunize experimental animal models. The results have shown that these proteins can enhance immunogenicity by generating an adequate immune response capable of protecting the model against an experimental infectious challenge, whereas protection is decreased when the specific antigen is administered without the Cry protein. Therefore, based on previous results and the structural homology between Cry proteins, these molecules have arisen as potential adjuvants in the development of vaccines for both animals and humans. Finally, a model of the interaction of Cry proteins with different components of the immune response is proposed.