KVFinder-web: a web-based application for detecting and characterizing biomolecular cavities.

Molecular interactions that modulate catalytic processes occur mainly in cavities throughout the molecular surface. Such interactions occur with specific small molecules due to geometric and physicochemical complementarity with the receptor. In this scenario, we present KVFinder-web, an open-source web-based application of parKVFinder software for cavity detection and characterization of biomolecular structures. The KVFinder-web has two independent components: a RESTful web service and a web graphical portal. Our web service, KVFinder-web service, handles client requests, manages accepted jobs, and performs cavity detection and characterization on accepted jobs. Our graphical web portal, KVFinder-web portal, provides a simple and straightforward page for cavity analysis, which customizes detection parameters, submits jobs to the web service component, and displays cavities and characterizations. We provide a publicly available KVFinder-web at https://kvfinder-web.cnpem.br, running in a cloud environment as docker containers. Further, this deployment type allows KVFinder-web components to be configured locally and customized according to user demand. Hence, users may run jobs on a locally configured service or our public KVFinder-web.

Mapping Conformational Changes in the Saliva Proteome Potentially Associated with Oral Cancer Aggressiveness.

Diverse proteomics-based strategies have been applied to saliva to quantitatively identify diagnostic and prognostic targets for oral cancer. Considering that these targets may be regulated by events that do not imply variation in protein abundance levels, we hypothesized that changes in protein conformation can be associated with diagnosis and prognosis, revealing biological processes and novel targets of clinical relevance. For this, we employed limited proteolysis-mass spectrometry in saliva samples to explore structural alterations, comparing the proteome of healthy control and oral squamous cell carcinoma (OSCC) patients with and without lymph node metastasis. Thirty-six proteins with potential structural rearrangements were associated with clinical patient features including transketolase and its interacting partners. Moreover, N-glycosylated peptides contribute to structural rearrangements of potential diagnostic and prognostic markers. Altogether, this approach utilizes saliva proteins to search for targets for diagnosing and prognosing oral cancer and can guide the discovery of potential regulated sites beyond protein-level abundance.

Habitat and Host Species Drive the Structure of Bacterial Communities of Two Neotropical Trap-Jaw Odontomachus Ants : Habitat and Host Species Drive the Structure of Bacterial Communities of Two Neotropical Trap-Jaw Odontomachus Ants.

Ants have long been known for their associations with other taxa, including macroscopic fungi and symbiotic bacteria. Recently, many ant species have had the composition and function of their bacterial communities investigated. Due to its behavioral and ecological diversity, the subfamily Ponerinae deserves more attention regarding its associated microbiota. Here, we used the V4 region of the 16S rRNA gene to characterize the bacterial communities of Odontomachus chelifer (ground-nesting) and Odontomachus hastatus (arboreal), two ponerine trap-jaw species commonly found in the Brazilian savanna ("Cerrado") and Atlantic rainforest. We investigated habitat effects (O. chelifer in the Cerrado and the Atlantic rainforest) and species-specific effects (both species in the Atlantic rainforest) on the bacterial communities' structure (composition and abundance) in two different body parts: cuticle and gaster. Bacterial communities differed in all populations studied. Cuticular communities were more diverse, while gaster communities presented variants common to other ants, including Wolbachia and Candidatus Tokpelaia hoelldoblerii. Odontomachus chelifer populations presented different communities in both body parts, highlighting the influence of habitat type. In the Atlantic rainforest, the outcome depended on the body part targeted. Cuticular communities were similar between species, reinforcing the habitat effect on bacterial communities, which are mainly composed of environmentally acquired taxa. Gaster communities, however, differed between the two Odontomachus species, suggesting species-specific effects and selective filters. Unclassified Firmicutes and uncultured Rhizobiales variants are the main components accounting for the observed differences. Our study indicates that both host species and habitat act synergistically, but to different degrees, to shape the bacterial communities in these Odontomachus species.

Cavity Characterization in Supramolecular Cages.

Confining molecular guests within artificial hosts has provided a major driving force in the rational design of supramolecular cages with tailored properties. Over the last 30 years, a set of design strategies have been developed that enabled the controlled synthesis of a myriad of cages. Recently, there has been a growing interest in involving in silico methods in this toolbox. Cavity shape and size are important parameters that can be easily accessed by inexpensive geometric algorithms. Although these algorithms are well developed for the detection of nonartificial cavities (e.g., enzymes), they are not routinely used for the rational design of supramolecular cages. In order to test the capabilities of this tool, we have evaluated the performance and characteristics of seven different cavity characterization software in the context of 22 analogues of well-known supramolecular cages. Among the tested software, KVFinder project and Fpocket proved to be the most software to characterize supramolecular cavities. With the results of this work, we aim to popularize this underused technique within the supramolecular community.

Microsatellites for the Neotropical Ant, Odontomachus chelifer (Hymenoptera: Formicidae).

Odontomachus chelifer (Latreille) (Ponerinae) is a ground-dwelling, predominantly carnivorous ant whose colonies may contain multiple egg-laying queens and are potentially susceptible to border effects in the Brazilian savanna known as Cerrado. The ecology and natural history of O. chelifer is well studied, but very little is known about the genetic diversity of O. chelifer colonies. In this study, we developed microsatellite markers for the study of genetic variation in O. chelifer. We created a microsatellite-enriched library that resulted in the development and characterization of 22 markers, of which 18 were found to be polymorphic in the population studied. The mean expected heterozygosity was 0.59, whereas the mean rarified allelic richness was determined as 4.27 alleles per locus. The polymorphism level detected was similar to genetic diversity estimates found in other poneromorph ant species. The microsatellites developed here are likely to be useful for the investigation of colony structure, functional polygyny, breeding system, and population genetics in O. chelifer. Moreover, the description of O. chelifer's genetic diversity is crucial for its conservation and maintenance of its ecological role in the Cerrado savanna.

Calcium and magnesium ions modulate the oligomeric state and function of mitochondrial 2-Cys peroxiredoxins in Leishmania parasites.

Leishmania parasites have evolved a number of strategies to cope with the harsh environmental changes during mammalian infection. One of these mechanisms involves the functional gain that allows mitochondrial 2-Cys peroxiredoxins to act as molecular chaperones when forming decamers. This function is critical for parasite infectivity in mammals, and its activation has been considered to be controlled exclusively by the enzyme redox state under physiological conditions. Herein, we have revealed that magnesium and calcium ions play a major role in modulating the ability of these enzymes to act as molecular chaperones, surpassing the redox effect. These ions are directly involved in mitochondrial metabolism and participate in a novel mechanism to stabilize the decameric form of 2-Cys peroxiredoxins in Leishmania mitochondria. Moreover, we have demonstrated that a constitutively dimeric Prx1m mutant impairs the survival of Leishmania under heat stress, supporting the central role of the chaperone function of Prx1m for Leishmania parasites during the transition from insect to mammalian hosts.

Indirect effects of mutualism: ant-treehopper associations deter pollinators and reduce reproduction in a tropical shrub.

Animal-pollinated plants can be susceptible to changes in pollinator availability. Honeydew-producing treehoppers frequently occur on inflorescences, potentially enhancing ant-mediated negative effects on pollination services. However, the effect of ant-attended, honeydew-producing insects on plant reproduction remains uncertain. We recorded the abundance of treehoppers and ants on Byrsonima intermedia (Malpighiaceae), and monitored floral visitors in a Brazilian cerrado savanna. We manipulated the presence of ants and ant-treehopper associations on inflorescences to assess their effect on pollination and fruit formation. We used dried ants pinned to inflorescences to evaluate the effect of ant presence and ant identity on potential pollinators. Results show that the presence of treehoppers increases ant abundance on flowers and disrupts pollination by oil-collecting bees, decreasing the frequency and duration of floral visits and reducing fruit and seed set. Treehopper herbivory has no direct effect on fruit or seed production, which are independent of treehopper density. Pinned ants promote avoidance by floral visitors, reducing the number of visits. Ant identity mediates visitation decisions, with Ectatomma brunneum causing greater avoidance by floral visitors than Camponotus rufipes. Field videos show that pollinating bees are harassed by ants near flowers, prompting avoidance behavior by the bees. This is the first demonstration of indirect effects by honeydew-gathering ants, via disrupted pollination, on plant reproduction in tropical cerrado savanna. Our results highlight the importance of studying other interactions near flowers, in addition to just observing pollinators, for a proper understanding of plant reproduction.

Fear Mediates Trophic Cascades: Nonconsumptive Effects of Predators Drive Aquatic Ecosystem Function.

Predators control prey populations and influence communities and the functioning of ecosystems through a combination of consumptive and nonconsumptive effects. These effects can be locally confined to one ecosystem but can also be extended to neighboring ecosystems. In this study, we investigated the nonconsumptive effects of terrestrial avian predators on the communities of aquatic invertebrates inhabiting bromeliads and on the functioning of these natural ecosystems. Bromeliads with stuffed birds placed nearby showed a decrease in aquatic damselfly larvae abundance and biomass, and we can infer that these changes were caused by antipredator responses. These larvae, which are top predators in bromeliad ecosystems, changed the composition of the entire aquatic invertebrate community. While total species richness, mesopredator richness, and shredder abundance increased in the presence of birds, scraper biomass decreased, possibly as a consequence of the increase in mesopredator richness. High scraper biomass in the absence of birds may have accelerated detrital decomposition, making more nutrients available for bromeliads, which grew more. These results show that nonconsumptive effects triggered by terrestrial predators can cascade down to lower trophic levels and dramatically affect the functioning of aquatic ecosystems, which can in turn alter nutrient provision to terrestrial ecosystems.

Effects of predatory ants within and across ecosystems in bromeliad food webs.

Predation is one of the most fundamental ecological processes affecting biotic communities. Terrestrial predators that live at ecosystem boundaries may alter the diversity of terrestrial organisms, but they may also have cross-ecosystem cascading effects when they feed on organisms with complex life cycles (i.e. organisms that shift from aquatic juvenile stages to terrestrial adult stages) or inhibit female oviposition in the aquatic environment. The predatory ant Odontomachus hastatus establishes its colonies among roots of Vriesea procera, an epiphytic bromeliad species with water-filled tanks that shelters many terrestrial and aquatic organisms. Ants may impact terrestrial communities and deter adult insects from ovipositing in the water of bromeliads via consumptive and non-consumptive effects. Ants do not forage within the aquatic environment; thus, they may be more efficient predators on terrestrial organisms. Therefore, we predict that ants will have stronger effects on terrestrial than aquatic food webs. However, such effects may also be site contingent and depend on the local composition of food webs. To test our hypothesis, we surveyed bromeliads with and without O. hastatus colonies from three different coastal field sites in the Atlantic Forest of southeast Brazil, and quantified the effect of this predatory ant on the composition, density and richness of aquatic and terrestrial metazoans found in these bromeliads. We found that ants changed the composition and reduced the overall density of aquatic and terrestrial metazoans in bromeliad ecosystems. However, effects of ants on species diversity were contingent on site. In general terms, the effects of the ant on aquatic and terrestrial metazoan communities were similar in strength and magnitude. Ants reduced the density of virtually all aquatic functional groups, especially detritivore insects as well as metazoans that reach bromeliads through phoresy on the skin of terrestrial animals (i.e. Ostracoda and Helobdella sp.). Our results suggest that the cross-ecosystem effect of this terrestrial predator on the aquatic metazoans was at least as strong as its within-ecosystem effect on the terrestrial ecosystem, and demonstrates that the same predator can simultaneously initiate cascades in multiple ecosystems.

Mutualism exploitation: predatory drosophilid larvae sugar-trap ants and jeopardize facultative ant-plant mutualism.

An open question in the evolutionary ecology of ant-plant facultative mutualism is how other members of the associated community can affect the interaction to a point where reciprocal benefits are disrupted. While visiting Qualea grandiflora shrubs to collect sugary rewards at extrafloral nectaries, tropical savanna ants deter herbivores and reduce leaf damage. Here we show that larvae of the fly Rhinoleucophenga myrmecophaga, which develop on extrafloral nectaries, lure potentially mutualistic, nectar-feeding ants and prey on them. Foraging ants spend less time on fly-infested foliage. Field experiments showed that predation (or the threat of predation) on ants by fly larvae produces cascading effects through three trophic levels, resulting in fewer protective ants on leaves, increased numbers of chewing herbivores, and greater leaf damage. These results reveal an undocumented mode of mutualism exploitation by an opportunistic predator at a plant-provided food source, jeopardizing ant-derived protection services to the plant. Our study documents a rather unusual case of predation of adult ants by a dipteran species and demonstrates a top-down trophic cascade within a generalized ant-plant mutualism.

A multiobjective approach to the genetic code adaptability problem.

BACKGROUND: The organization of the canonical code has intrigued researches since it was first described. If we consider all codes mapping the 64 codes into 20 amino acids and one stop codon, there are more than 1.51×10(84) possible genetic codes. The main question related to the organization of the genetic code is why exactly the canonical code was selected among this huge number of possible genetic codes. Many researchers argue that the organization of the canonical code is a product of natural selection and that the code's robustness against mutations would support this hypothesis. In order to investigate the natural selection hypothesis, some researches employ optimization algorithms to identify regions of the genetic code space where best codes, according to a given evaluation function, can be found (engineering approach). The optimization process uses only one objective to evaluate the codes, generally based on the robustness for an amino acid property. Only one objective is also employed in the statistical approach for the comparison of the canonical code with random codes. We propose a multiobjective approach where two or more objectives are considered simultaneously to evaluate the genetic codes. RESULTS: In order to test our hypothesis that the multiobjective approach is useful for the analysis of the genetic code adaptability, we implemented a multiobjective optimization algorithm where two objectives are simultaneously optimized. Using as objectives the robustness against mutation with the amino acids properties polar requirement (objective 1) and robustness with respect to hydropathy index or molecular volume (objective 2), we found solutions closer to the canonical genetic code in terms of robustness, when compared with the results using only one objective reported by other authors. CONCLUSIONS: Using more objectives, more optimal solutions are obtained and, as a consequence, more information can be used to investigate the adaptability of the genetic code. The multiobjective approach is also more natural, because more than one objective was adapted during the evolutionary process of the canonical genetic code. Our results suggest that the evaluation function employed to compare genetic codes should consider simultaneously more than one objective, in contrast to what has been done in the literature.

Mechanistic strategies for catalysis adopted by evolutionary distinct family 43 arabinanases.

Arabinanases (ABNs, EC 3.2.1.99) are promising catalysts for environmentally friendly biomass conversion into energy and chemicals. These enzymes catalyze the hydrolysis of the α-1,5-linked L-arabinofuranoside backbone of plant cell wall arabinans releasing arabino-oligosaccharides and arabinose, the second most abundant pentose in nature. In this work, new findings about the molecular mechanisms governing activation, functional differentiation, and catalysis of GH43 ABNs are presented. Biophysical, mutational, and biochemical studies with the hyperthermostable two-domain endo-acting ABN from Thermotoga petrophila (TpABN) revealed how some GH43 ABNs are activated by calcium ions via hyperpolarization of the catalytically relevant histidine and the importance of the ancillary domain for catalysis and conformational stability. On the other hand, the two GH43 ABNs from rumen metagenome, ARN2 and ARN3, presented a calcium-independent mechanism in which sodium is the most likely substituent for calcium ions. The crystal structure of the two-domain endo-acting ARN2 showed that its ability to efficiently degrade branched substrates is due to a larger catalytic interface with higher accessibility than that observed in other ABNs with preference for linear arabinan. Moreover, crystallographic characterization of the single-domain exo-acting ARN3 indicated that its cleavage pattern producing arabinose is associated with the chemical recognition of the reducing end of the substrate imposed by steric impediments at the aglycone-binding site. By structure-guided rational design, ARN3 was converted into a classical endo enzyme, confirming the role of the extended Arg(203)-Ala(230) loop in determining its action mode. These results reveal novel molecular aspects concerning the functioning of GH43 ABNs and provide new strategies for arabinan degradation.

P-I class metalloproteinase from Bothrops moojeni venom is a post-proline cleaving peptidase with kininogenase activity: insights into substrate selectivity and kinetic behavior.

Snake venom metalloproteinases (SVMPs) belonging to P-I class are able to hydrolyze extracellular matrix proteins and coagulation factors triggering local and systemic reactions by multiple molecular mechanisms that are not fully understood. BmooMPα-I, a P-I class SMVP from Bothrops moojeni venom, was active upon neuro- and vaso-active peptides including angiotensin I, bradykinin, neurotensin, oxytocin and substance P. Interestingly, BmooMPα-I showed a strong bias towards hydrolysis after proline residues, which is unusual for most of characterized peptidases. Moreover, the enzyme showed kininogenase activity similar to that observed in plasma and cells by kallikrein. FRET peptide assays indicated a relative promiscuity at its S2-S'2 subsites, with proline determining the scissile bond. This unusual post-proline cleaving activity was confirmed by the efficient hydrolysis of the synthetic combinatorial library MCA-GXXPXXQ-EDDnp, described as resistant for canonical peptidases, only after Pro residues. Structural analysis of the tripeptide LPL complexed with BmooMPα-I, generated by molecular dynamics simulations, assisted in defining the subsites and provided the structural basis for subsite preferences such as the restriction of basic residues at the S2 subsite due to repulsive electrostatic effects and the steric impediment for large aliphatic or aromatic side chains at the S1 subsite. These new functional and structural findings provided a further understanding of the molecular mechanisms governing the physiological effects of this important class of enzymes in envenomation process.

Ant-caterpillar antagonism at the community level: interhabitat variation of tritrophic interactions in a neotropical savanna.

Ant foraging on foliage can substantially affect how phytophagous insects use host plants and represents a high predation risk for caterpillars, which are important folivores. Ant-plant-herbivore interactions are especially pervasive in cerrado savanna due to continuous ant visitation to liquid food sources on foliage (extrafloral nectaries, insect honeydew). While searching for liquid rewards on plants, aggressive ants frequently attack or kill insect herbivores, decreasing their numbers. Because ants vary in diet and aggressiveness, their effect on herbivores also varies. Additionally, the differential occurrence of ant attractants (plant and insect exudates) on foliage produces variable levels of ant foraging within local floras and among localities. Here, we investigate how variation of ant communities and of traits among host plant species (presence or absence of ant attractants) can change the effect of carnivores (predatory ants) on herbivore communities (caterpillars) in a cerrado savanna landscape. We sampled caterpillars and foliage-foraging ants in four cerrado localities (70-460 km apart). We found that: (i) caterpillar infestation was negatively related with ant visitation to plants; (ii) this relationship depended on local ant abundance and species composition, and on local preference by ants for plants with liquid attractants; (iii) this was not related to local plant richness or plant size; (iv) the relationship between the presence of ant attractants and caterpillar abundance varied among sites from negative to neutral; and (v) caterpillars feeding on plants with ant attractants are more resistant to ant predation than those feeding on plants lacking attractants. Liquid food on foliage mediates host plant quality for lepidopterans by promoting generalized ant-caterpillar antagonism. Our study in cerrado shows that the negative effects of generalist predatory ants on herbivores are detectable at a community level, affecting patterns of abundance and host plant use by lepidopterans. The magnitude of ant-induced effects on caterpillar occurrence across the cerrado landscape may depend on how ants use plants locally and how they respond to liquid food on plants at different habitats. This study enhances the relevance of plant-ant and ant-herbivore interactions in cerrado and highlights the importance of a tritrophic perspective in this ant-rich environment.

KVFinder: steered identification of protein cavities as a PyMOL plugin.

BACKGROUND: The characterization of protein binding sites is a major challenge in computational biology. Proteins interact with a wide variety of molecules and understanding of such complex interactions is essential to gain deeper knowledge of protein function. Shape complementarity is known to be important in determining protein-ligand interactions. Furthermore, these protein structural features have been shown to be useful in assisting medicinal chemists during lead discovery and optimization. RESULTS: We developed KVFinder, a highly versatile and easy-to-use tool for cavity prospection and spatial characterization. KVFinder is a geometry-based method that has an innovative customization of the search space. This feature provides the possibility of cavity segmentation, which alongside with the large set of customizable parameters, allows detailed cavity analyses. Although the main focus of KVFinder is the steered prospection of cavities, we tested it against a benchmark dataset of 198 known drug targets in order to validate our software and compare it with some of the largely accepted methods. Using the one click mode, we performed better than most of the other methods, staying behind only of hybrid prospection methods. When using just one of KVFinder's customizable features, we were able to outperform all other compared methods. KVFinder is also user friendly, as it is available as a PyMOL plugin, or command-line version. CONCLUSION: KVFinder presents novel usability features, granting full customizable and highly detailed cavity prospection on proteins, alongside with a friendly graphical interface. KVFinder is freely available on http://lnbio.cnpem.br/bioinformatics/main/software/.

Structural insights into functional overlapping and differentiation among myosin V motors.

Myosin V (MyoV) motors have been implicated in the intracellular transport of diverse cargoes including vesicles, organelles, RNA-protein complexes, and regulatory proteins. Here, we have solved the cargo-binding domain (CBD) structures of the three human MyoV paralogs (Va, Vb, and Vc), revealing subtle structural changes that drive functional differentiation and a novel redox mechanism controlling the CBD dimerization process, which is unique for the MyoVc subclass. Moreover, the cargo- and motor-binding sites were structurally assigned, indicating the conservation of residues involved in the recognition of adaptors for peroxisome transport and providing high resolution insights into motor domain inhibition by CBD. These results contribute to understanding the structural requirements for cargo transport, autoinhibition, and regulatory mechanisms in myosin V motors.

Structural shifts of aldehyde dehydrogenase enzymes were instrumental for the early evolution of retinoid-dependent axial patterning in metazoans.

Aldehyde dehydrogenases (ALDHs) catabolize toxic aldehydes and process the vitamin A-derived retinaldehyde into retinoic acid (RA), a small diffusible molecule and a pivotal chordate morphogen. In this study, we combine phylogenetic, structural, genomic, and developmental gene expression analyses to examine the evolutionary origins of ALDH substrate preference. Structural modeling reveals that processing of small aldehydes, such as acetaldehyde, by ALDH2, versus large aldehydes, including retinaldehyde, by ALDH1A is associated with small versus large substrate entry channels (SECs), respectively. Moreover, we show that metazoan ALDH1s and ALDH2s are members of a single ALDH1/2 clade and that during evolution, eukaryote ALDH1/2s often switched between large and small SECs after gene duplication, transforming constricted channels into wide opened ones and vice versa. Ancestral sequence reconstructions suggest that during the evolutionary emergence of RA signaling, the ancestral, narrow-channeled metazoan ALDH1/2 gave rise to large ALDH1 channels capable of accommodating bulky aldehydes, such as retinaldehyde, supporting the view that retinoid-dependent signaling arose from ancestral cellular detoxification mechanisms. Our analyses also indicate that, on a more restricted evolutionary scale, ALDH1 duplicates from invertebrate chordates (amphioxus and ascidian tunicates) underwent switches to smaller and narrower SECs. When combined with alterations in gene expression, these switches led to neofunctionalization from ALDH1-like roles in embryonic patterning to systemic, ALDH2-like roles, suggesting functional shifts from signaling to detoxification.

Exploring the Potential of Structure-Based Deep Learning Approaches for T cell Receptor Design.

Deep learning methods, trained on the increasing set of available protein 3D structures and sequences, have substantially impacted the protein modeling and design field. These advancements have facilitated the creation of novel proteins, or the optimization of existing ones designed for specific functions, such as binding a target protein. Despite the demonstrated potential of such approaches in designing general protein binders, their application in designing immunotherapeutics remains relatively unexplored. A relevant application is the design of T cell receptors (TCRs). Given the crucial role of T cells in mediating immune responses, redirecting these cells to tumor or infected target cells through the engineering of TCRs has shown promising results in treating diseases, especially cancer. However, the computational design of TCR interactions presents challenges for current physics-based methods, particularly due to the unique natural characteristics of these interfaces, such as low affinity and cross-reactivity. For this reason, in this study, we explored the potential of two structure-based deep learning protein design methods, ProteinMPNN and ESM-IF, in designing fixed-backbone TCRs for binding target antigenic peptides presented by the MHC through different design scenarios. To evaluate TCR designs, we employed a comprehensive set of sequence- and structure-based metrics, highlighting the benefits of these methods in comparison to classical physics-based design methods and identifying deficiencies for improvement.

Identification of novel interaction between ADAM17 (a disintegrin and metalloprotease 17) and thioredoxin-1.

ADAM17, which is also known as TNFα-converting enzyme, is the major sheddase for the EGF receptor ligands and is considered to be one of the main proteases responsible for the ectodomain shedding of surface proteins. How a membrane-anchored proteinase with an extracellular catalytic domain can be activated by inside-out regulation is not completely understood. We characterized thioredoxin-1 (Trx-1) as a partner of the ADAM17 cytoplasmic domain that could be involved in the regulation of ADAM17 activity. We induced the overexpression of the ADAM17 cytoplasmic domain in HEK293 cells, and ligands able to bind this domain were identified by MS after protein immunoprecipitation. Trx-1 was also validated as a ligand of the ADAM17 cytoplasmic domain and full-length ADAM17 recombinant proteins by immunoblotting, immunolocalization, and solid phase binding assay. In addition, using nuclear magnetic resonance, it was shown in vitro that the titration of the ADAM17 cytoplasmic domain promotes changes in the conformation of Trx-1. The MS analysis of the cross-linked complexes showed cross-linking between the two proteins by lysine residues. To further evaluate the functional role of Trx-1, we used a heparin-binding EGF shedding cell model and observed that the overexpression of Trx-1 in HEK293 cells could decrease the activity of ADAM17, activated by either phorbol 12-myristate 13-acetate or EGF. This study identifies Trx-1 as a novel interaction partner of the ADAM17 cytoplasmic domain and suggests that Trx-1 is a potential candidate that could be involved in ADAM17 activity regulation.

Functional diversification of cerato-platanins in Moniliophthora perniciosa as seen by differential expression and protein function specialization.

Cerato-platanins (CP) are small, cysteine-rich fungal-secreted proteins involved in the various stages of the host-fungus interaction process, acting as phytotoxins, elicitors, and allergens. We identified 12 CP genes (MpCP1 to MpCP12) in the genome of Moniliophthora perniciosa, the causal agent of witches' broom disease in cacao, and showed that they present distinct expression profiles throughout fungal development and infection. We determined the X-ray crystal structures of MpCP1, MpCP2, MpCP3, and MpCP5, representative of different branches of a phylogenetic tree and expressed at different stages of the disease. Structure-based biochemistry, in combination with nuclear magnetic resonance and mass spectrometry, allowed us to define specialized capabilities regarding self-assembling and the direct binding to chitin and N-acetylglucosamine (NAG) tetramers, a fungal cell wall building block, and to map a previously unknown binding region in MpCP5. Moreover, fibers of MpCP2 were shown to act as expansin and facilitate basidiospore germination whereas soluble MpCP5 blocked NAG6-induced defense response. The correlation between these roles, the fungus life cycle, and its tug-of-war interaction with cacao plants is discussed.