Predictors of standardized in-service examination performance and residency outcomes in a graduate periodontics program.

PURPOSE/OBJECTIVES: The objectives of this study were to assess the influence of learner- and education-related factors on standardized in-service examination performance and determine whether in-service examination scores predict residency outcomes. METHODS: American Academy of Periodontology (AAP) In-service Examination (AIE) scores from 10 periodontics residency classes at a single center were recorded and compared against a panel of learner- and education-related variables using multiple linear regression models. Defined residency outcome measures were analyzed against AIE scores using binomial logistic regression. RESULTS: No evaluated learner- or education-related variable was a statistically significant predictor of AIE score in this study sample. Likewise, AIE score was not a statistically significant predictor of any assessed residency outcome. CONCLUSIONS: The AAP has performed a tremendous service to periodontics residents and programs by marshaling the leadership and expertise necessary to offer a professionally constructed assessment instrument. However, in the current study, no relationship could be identified between AIE score and any outcome, including first-attempt board certification. The AAP In-service Committee appears well situated to provide additional leadership focusing on exam implementation, which may enhance AIE value in competency decision making.

Fragment-Based Ab Initio Phasing of Peptidic Nanocrystals by MicroED.

Electron diffraction (MicroED/3DED) can render the three-dimensional atomic structures of molecules from previously unamenable samples. The approach has been particularly transformative for peptidic structures, where MicroED has revealed novel structures of naturally occurring peptides, synthetic protein fragments, and peptide-based natural products. Despite its transformative potential, MicroED is beholden to the crystallographic phase problem, which challenges its de novo determination of structures. ARCIMBOLDO, an automated, fragment-based approach to structure determination, eliminates the need for atomic resolution, instead enforcing stereochemical constraints through libraries of small model fragments, and discerning congruent motifs in solution space to ensure validation. This approach expands the reach of MicroED to presently inaccessible peptide structures including fragments of human amyloids, and yeast and mammalian prions. For electron diffraction, fragment-based phasing portends a more general phasing solution with limited model bias for a wider set of chemical structures.

Likelihood-based signal and noise analysis for docking of models into cryo-EM maps.

Fast, reliable docking of models into cryo-EM maps requires understanding of the errors in the maps and the models. Likelihood-based approaches to errors have proven to be powerful and adaptable in experimental structural biology, finding applications in both crystallography and cryo-EM. Indeed, previous crystallographic work on the errors in structural models is directly applicable to likelihood targets in cryo-EM. Likelihood targets in Fourier space are derived here to characterize, based on the comparison of half-maps, the direction- and resolution-dependent variation in the strength of both signal and noise in the data. Because the signal depends on local features, the signal and noise are analysed in local regions of the cryo-EM reconstruction. The likelihood analysis extends to prediction of the signal that will be achieved in any docking calculation for a model of specified quality and completeness. A related calculation generalizes a previous measure of the information gained by making the cryo-EM reconstruction.

Likelihood-based docking of models into cryo-EM maps.

Optimized docking of models into cryo-EM maps requires exploiting an understanding of the signal expected in the data to minimize the calculation time while maintaining sufficient signal. The likelihood-based rotation function used in crystallography can be employed to establish plausible orientations in a docking search. A phased likelihood translation function yields scores for the placement and rigid-body refinement of oriented models. Optimized strategies for choices of the resolution of data from the cryo-EM maps to use in the calculations and the size of search volumes are based on expected log-likelihood-gain scores computed in advance of the search calculation. Tests demonstrate that the new procedure is fast, robust and effective at placing models into even challenging cryo-EM maps.

Putting AlphaFold models to work with phenix.process_predicted_model and ISOLDE.

AlphaFold has recently become an important tool in providing models for experimental structure determination by X-ray crystallography and cryo-EM. Large parts of the predicted models typically approach the accuracy of experimentally determined structures, although there are frequently local errors and errors in the relative orientations of domains. Importantly, residues in the model of a protein predicted by AlphaFold are tagged with a predicted local distance difference test score, informing users about which regions of the structure are predicted with less confidence. AlphaFold also produces a predicted aligned error matrix indicating its confidence in the relative positions of each pair of residues in the predicted model. The phenix.process_predicted_model tool downweights or removes low-confidence residues and can break a model into confidently predicted domains in preparation for molecular replacement or cryo-EM docking. These confidence metrics are further used in ISOLDE to weight torsion and atom-atom distance restraints, allowing the complete AlphaFold model to be interactively rearranged to match the docked fragments and reducing the need for the rebuilding of connecting regions.

Improved AlphaFold modeling with implicit experimental information.

Machine-learning prediction algorithms such as AlphaFold and RoseTTAFold can create remarkably accurate protein models, but these models usually have some regions that are predicted with low confidence or poor accuracy. We hypothesized that by implicitly including new experimental information such as a density map, a greater portion of a model could be predicted accurately, and that this might synergistically improve parts of the model that were not fully addressed by either machine learning or experiment alone. An iterative procedure was developed in which AlphaFold models are automatically rebuilt on the basis of experimental density maps and the rebuilt models are used as templates in new AlphaFold predictions. We show that including experimental information improves prediction beyond the improvement obtained with simple rebuilding guided by the experimental data. This procedure for AlphaFold modeling with density has been incorporated into an automated procedure for interpretation of crystallographic and electron cryo-microscopy maps.

Assessing the utility of CASP14 models for molecular replacement.

The assessment of CASP models for utility in molecular replacement is a measure of their use in a valuable real-world application. In CASP7, the metric for molecular replacement assessment involved full likelihood-based molecular replacement searches; however, this restricted the assessable targets to crystal structures with only one copy of the target in the asymmetric unit, and to those where the search found the correct pose. In CASP10, full molecular replacement searches were replaced by likelihood-based rigid-body refinement of models superimposed on the target using the LGA algorithm, with the metric being the refined log-likelihood-gain (LLG) score. This enabled multi-copy targets and very poor models to be evaluated, but a significant further issue remained: the requirement of diffraction data for assessment. We introduce here the relative-expected-LLG (reLLG), which is independent of diffraction data. This reLLG is also independent of any crystal form, and can be calculated regardless of the source of the target, be it X-ray, NMR or cryo-EM. We calibrate the reLLG against the LLG for targets in CASP14, showing that it is a robust measure of both model and group ranking. Like the LLG, the reLLG shows that accurate coordinate error estimates add substantial value to predicted models. We find that refinement by CASP groups can often convert an inadequate initial model into a successful MR search model. Consistent with findings from others, we show that the AlphaFold2 models are sufficiently good, and reliably so, to surpass other current model generation strategies for attempting molecular replacement phasing.

Accurate prediction of protein structures and interactions using a three-track neural network.

DeepMind presented notably accurate predictions at the recent 14th Critical Assessment of Structure Prediction (CASP14) conference. We explored network architectures that incorporate related ideas and obtained the best performance with a three-track network in which information at the one-dimensional (1D) sequence level, the 2D distance map level, and the 3D coordinate level is successively transformed and integrated. The three-track network produces structure predictions with accuracies approaching those of DeepMind in CASP14, enables the rapid solution of challenging x-ray crystallography and cryo-electron microscopy structure modeling problems, and provides insights into the functions of proteins of currently unknown structure. The network also enables rapid generation of accurate protein-protein complex models from sequence information alone, short-circuiting traditional approaches that require modeling of individual subunits followed by docking. We make the method available to the scientific community to speed biological research.

Structure of the class XI myosin globular tail reveals evolutionary hallmarks for cargo recognition in plants.

The plant-specific class XI myosins (MyoXIs) play key roles at the molecular, cellular and tissue levels, engaging diverse adaptor proteins to transport cargoes along actin filaments. To recognize their cargoes, MyoXIs have a C-terminal globular tail domain (GTD) that is evolutionarily related to those of class V myosins (MyoVs) from animals and fungi. Despite recent advances in understanding the functional roles played by MyoXI in plants, the structure of its GTD, and therefore the molecular determinants for cargo selectivity and recognition, remain elusive. In this study, the first crystal structure of a MyoXI GTD, that of MyoXI-K from Arabidopsis thaliana, was elucidated at 2.35 Šresolution using a low-identity and fragment-based phasing approach in ARCIMBOLDO_SHREDDER. The results reveal that both the composition and the length of the α5-α6 loop are distinctive features of MyoXI-K, providing evidence for a structural stabilizing role for this loop, which is otherwise carried out by a molecular zipper in MyoV GTDs. The crystal structure also shows that most of the characterized cargo-binding sites in MyoVs are not conserved in plant MyoXIs, pointing to plant-specific cargo-recognition mechanisms. Notably, the main elements involved in the self-regulation mechanism of MyoVs are conserved in plant MyoXIs, indicating this to be an ancient ancestral trait.

Arbovirus en Uruguay, un problema potencial. Revisión desde una óptica perinatal / Arbovirus in Uruguay, a potential problem. Review from a perinatal perspective / Arbovírus no Uruguai, um problema potencial. Revisão a partir de uma perspectiva perinatal

Resumen: Los arbovirus son microorganismos transmitidos al ser humano por artrópodos. Existen más de 100 tipos de arbovirus, con una presentación clínica común para todos ellos de fiebre, síntomas articulares, hemorrágicos y neurológicos1. Además, representan un riesgo particular para la mujer embarazada y el feto por el potencial teratogénico que algunos de ellos presentan, particularmente a nivel del sistema nervioso central. Utilizando los motores de búsqueda de Pubmed y de Google Scholar, realizamos una búsqueda bibliográfica enfocada a los arbovirus en general y luego enfocada en cada una de las patologías específicas abordadas, que fueron definidas en función del riesgo de ingreso a nuestro país y sus potenciales consecuencias. El objetivo de esta revisión es analizar las principales características de presentación de los arbovirus, en particular dengue, zika, chikungunya y fiebre amarilla ante la eventualidad de la aparición de casos de transmisión vertical, para que nuestros recursos humanos especializados tengan un marco de referencia del manejo actualizado. Esta revisión nos permitió concluir sobre los elementos comunes de estas virosis, así como sus potenciales afectaciones en el feto y en el recién nacido, y nos planteó el desafío vinculado a su dificultad diagnóstica por las reacciones cruzadas.

Summary: Arbovirus are microorganisms transmitted to humans through arthropods. There are more than 100 different arboviruses with a common clinical presentation of fever, articular, hemorrhagic and neurological symptoms1. They represent a mayor risk to pregnant women and fetuses due to their teratogenic effect, particularly affecting the central nervous system. We performed a specific search focused on arbovirus using search engines Pubmed and Google Scholar and we classified evidence according to the risk of a certain virus entering our country and its potential consequences. The objective of this review is to update the main clinical characteristics of arbovirus, particularly Dengue, Zika, Chikungunya and Yellow Fever particularly due to the potential introduction of these viruses in our country and the possibility of vertical transmission, so that our human resources have a guide on how to approach these patients nowadays. This review allowed us to conclude on common characteristics of these viruses, their possible consequences on fetus and newborns, and concluding on the difficulty of etiological diagnosis due to cross reactions.

Resumo: Os arbovírus são microrganismos transmitidos ao homem por artrópodes. Existem mais de 100 tipos de arbovírus, com apresentação clínica comum a todos eles de febre, sintomas articulares, hemorrágicos e neurológicos1. Além disso, representam um risco particular para a gestante e para o feto devido ao potencial teratogênico que alguns deles apresentam, principalmente ao nível do sistema nervoso central. Realizamos uma busca bibliográfica utilizando Pubmed e Google Scholar sobre arbovírus em geral e posteriormente, sobre cada uma das patologias específicas abordadas que foram definidas de acordo com o risco de entrada em nosso país e suas possíveis consequências. O objetivo desta revisão é analisar as principais características de apresentação dos arbovírus, em particular dengue, Zika, Chikungunya e febre amarela, considerando a possibilidade de casos de transmissão vertical, para servir como uma referência atualizada para os profissionais especializados nesta área. Esta revisão permitiu elaborar conclusões sobre os elementos comuns a esses vírus, bem como suas possíveis afetações no feto e no recém-nascido, e apresentou o desafio relacionado à dificuldade diagnóstica devido às reações cruzadas.

Metastatic TNBC is closely associated with a fused mitochondrial morphology and a glycolytic and lipogenic metabolism.

Mitochondria modify their function and morphology to satisfy the bioenergetic demand of the cells. Cancer cells take advantage of these features to sustain their metabolic, proliferative, metastatic, and survival necessities. Understanding the morphological changes to mitochondria in the different grades of triple-negative breast cancer (TNBC) could help to design new treatments. Consequently, this research explored mitochondrial morphology and the gene expression of some proteins related to mitochondrial dynamics, as well as proteins associated with oxidative and non-oxidative metabolism in metastatic and non-metastatic TNBC. We found that mitochondrial morphology and metabolism are different in metastatic and non-metastatic TNBC. In metastatic TNBC, there is overexpression of genes related to mitochondrial dynamics, fatty-acid metabolism, and glycolysis. These features are accompanied by a fused mitochondrial morphology. By comparison, in non-metastatic TNBC, there is a stress-associated mitochondrial morphology with hyperfragmented mitochondria, accompanied by the upregulated expression of genes associated with the biogenesis of mitochondria; both of which are characteristics related to the higher production of reactive oxygen species observed in this cell line. These differences between metastatic and non-metastatic TNBC should provide a better understanding of metastasis and contribute to the development of improved specific and personalized therapies for TNBC.

Fragment-based determination of a proteinase K structure from MicroED data using ARCIMBOLDO_SHREDDER.

Structure determination of novel biological macromolecules by X-ray crystallography can be facilitated by the use of small structural fragments, some of only a few residues in length, as effective search models for molecular replacement to overcome the phase problem. Independence from the need for a complete pre-existing model with sequence similarity to the crystallized molecule is the primary appeal of ARCIMBOLDO, a suite of programs which employs this ab initio algorithm for phase determination. Here, the use of ARCIMBOLDO is investigated to overcome the phase problem with the electron cryomicroscopy (cryoEM) method known as microcrystal electron diffraction (MicroED). The results support the use of the ARCIMBOLDO_SHREDDER pipeline to provide phasing solutions for a structure of proteinase K from 1.6 Šresolution data using model fragments derived from the structures of proteins sharing a sequence identity of as low as 20%. ARCIMBOLDO_SHREDDER identified the most accurate polyalanine fragments from a set of distantly related sequence homologues. Alternatively, such templates were extracted in spherical volumes and given internal degrees of freedom to refine towards the target structure. Both modes relied on the rotation function in Phaser to identify or refine fragment models and its translation function to place them. Model completion from the placed fragments proceeded through phase combination of partial solutions and/or density modification and main-chain autotracing using SHELXE. The combined set of fragments was sufficient to arrive at a solution that resembled that determined by conventional molecular replacement using the known target structure as a search model. This approach obviates the need for a single, complete and highly accurate search model when phasing MicroED data, and permits the evaluation of large fragment libraries for this purpose.

ALEPH: a network-oriented approach for the generation of fragment-based libraries and for structure interpretation.

The analysis of large structural databases reveals general features and relationships among proteins, providing useful insight. A different approach is required to characterize ubiquitous secondary-structure elements, where flexibility is essential in order to capture small local differences. The ALEPH software is optimized for the analysis and the extraction of small protein folds by relying on their geometry rather than on their sequence. The annotation of the structural variability of a given fold provides valuable information for fragment-based molecular-replacement methods, in which testing alternative model hypotheses can succeed in solving difficult structures when no homology models are available or are successful. ARCIMBOLDO_BORGES combines the use of composite secondary-structure elements as a search model with density modification and tracing to reveal the rest of the structure when both steps are successful. This phasing method relies on general fold libraries describing variations around a given pattern of ß-sheets and helices extracted using ALEPH. The program introduces characteristic vectors defined from the main-chain atoms as a way to describe the geometrical properties of the structure. ALEPH encodes structural properties in a graph network, the exploration of which allows secondary-structure annotation, decomposition of a structure into small compact folds, generation of libraries of models representing a variation of a given fold and finally superposition of these folds onto a target structure. These functions are available through a graphical interface designed to interactively show the results of structure manipulation, annotation, fold decomposition, clustering and library generation. ALEPH can produce pictures of the graphs, structures and folds for publication purposes.

ALIXE: a phase-combination tool for fragment-based molecular replacement.

Fragment-based molecular replacement exploits the use of very accurate yet incomplete search models. In the case of the ARCIMBOLDO programs, consistent phase sets produced from the placement and refinement of fragments with Phaser can be combined in order to increase their signal before proceeding to the step of density modification and autotracing with SHELXE. The program ALIXE compares multiple phase sets, evaluating mean phase differences to determine their common origin, and subsequently produces sets of combined phases that group consistent solutions. In this work, its use on different scenarios of very partial molecular-replacement solutions and its performance after the development of a much-optimized set of algorithms are described. The program is available both standalone and integrated within the ARCIMBOLDO programs. ALIXE has been analysed to identify its rate-limiting steps while exploring the best parameterization to improve its performance and make this software efficient enough to work on modest hardware. The algorithm has been parallelized and redesigned to meet the typical landscape of solutions. Analysis of pairwise correlation between the phase sets has also been explored to test whether this would provide additional insight. ALIXE can be used to exhaustively analyse all partial solutions produced or to complement those already selected for expansion, and also to reduce the number of redundant solutions, which is particularly relevant to the case of coiled coils, or to combine partial solutions from different programs. In each case parallelization and optimization to provide speedup makes its use amenable to typical hardware found in crystallography. ARCIMBOLDO_BORGES and ARCIMBOLDO_SHREDDER now call on ALIXE by default.

SEQUENCE SLIDER: expanding polyalanine fragments for phasing with multiple side-chain hypotheses.

Fragment-based molecular-replacement methods can solve a macromolecular structure quasi-ab initio. ARCIMBOLDO, using a common secondary-structure or tertiary-structure template or a library of folds, locates these with Phaser and reveals the rest of the structure by density modification and autotracing in SHELXE. The latter stage is challenging when dealing with diffraction data at lower resolution, low solvent content, high ß-sheet composition or situations in which the initial fragments represent a low fraction of the total scattering or where their accuracy is low. SEQUENCE SLIDER aims to overcome these complications by extending the initial polyalanine fragment with side chains in a multisolution framework. Its use is illustrated on test cases and previously unknown structures. The selection and order of fragments to be extended follows the decrease in log-likelihood gain (LLG) calculated with Phaser upon the omission of each single fragment. When the starting substructure is derived from a remote homolog, sequence assignment to fragments is restricted by the original alignment. Otherwise, the secondary-structure prediction is matched to that found in fragments and traces. Sequence hypotheses are trialled in a brute-force approach through side-chain building and refinement. Scoring the refined models through their LLG in Phaser may allow discrimination of the correct sequence or filter the best partial structures for further density modification and autotracing. The default limits for the number of models to pursue are hardware dependent. In its most economic implementation, suitable for a single laptop, the main-chain trace is extended as polyserine rather than trialling models with different sequence assignments, which requires a grid or multicore machine. SEQUENCE SLIDER has been instrumental in solving two novel structures: that of MltC from 2.7 Šresolution data and that of a pneumococcal lipoprotein with 638 residues and 35% solvent content.

Tipificación de serotipos del virus del papiloma humano de alto riesgo / Typification of high-risk human papilomavirus serotypes

Resumen: OBJETIVO: Identificar los serotipos más frecuentes de virus del papiloma humano mediante pruebas al azar en pacientes previamente diagnosticadas con cáncer cervicouterino. MATERIALES Y MÉTODOS: Estudio prospectivo y observacional, efectuado en pacientes con displasia cervical, atendidas en el Hospital Regional Materno Infantil de Alta Especialidad de Nuevo León, Monterrey, entre enero y marzo de 2016. Criterios de inclusión: pacientes mayores de 18 años, que acudieron a la unidad médica para seguimiento y control ginecológico, con reporte anormal en la prueba de Papanicolaou, confirmado por histopatología, mediante biopsia dirigida por colposcopia. Criterios de exclusión: mujeres con histerectomía total por indicación de enfermedad benigna, sin antecedente de neoplasia intracervical; mayores a 70 años después de 3 citologías cervicales negativas en la década previa; pacientes que recibieron quimioterapia, radioterapia u otros tratamientos farmacológicos y quienes acudieron a revisión médica durante su ciclo menstrual. Para el análisis de los datos se utilizó estadística descriptiva. RESULTADOS: Se registraron 30 pacientes. Las clasificaciones más frecuentes de neoplasia cervical fueron: NIC-1 (n = 15), NIC-2 (n = 9) y NIC-3 (n = 6). Todas las pacientes analizadas tuvieron, al menos, un serotipo de VPH de alto riesgo. Los serotipos identificados con mayor frecuencia fueron el 31 y 33 (n = 18). En 18 pacientes se encontraron 6 o más serotipos de VPH. De 15 pacientes con lesiones de alto grado, 8 tuvieron la asociación de serotipos 31 y 33, y en 6 se identificó un serotipo aislado (16 y 51). CONCLUSIONES: Los serotipos identificados con mayor frecuencia fueron el 31 y 33. Desafortunadamente, la vacuna nonavalente que protege contra los serotipos más frecuentes de VPH no se encuentra disponible en Latinoamérica.

Abstract: OBJECTIVE: To identify the most frequent serotypes of human papillomavirus through random testing of patients previously diagnosed with cervical cancer. MATERIALS AND METHODS: A prospective, observational study carried out in patients with cervical dysplasia, treated at the High Specialty Regional Maternal and Child Hospital of Nuevo León, Monterrey. Inclusion criteria: patients over 18 years of age, who attended the Dysplasia Clinic of the High Specialty Regional Maternal and Child Hospital for gynecological follow-up and control, with an abnormal result in the Papanicolaou test, confirmed by histopathology, by means of colposcopy-directed biopsy. Before the procedures (cervical cytology), Exclusion criteria: women with total hysterectomy due to indications of benign disease, without a history of intracervical neoplasia; older than 70 years after 3 negative cervical cytology in the previous decade; patients who received chemotherapy, radiotherapy or other pharmacological treatments and who received medical check-ups during their menstrual cycle. Descriptive statistics were used for data analysis. RESULTS: 30 patients were registered. The most frequent classification of cervical neoplasia was: CIN1 (n = 15), CIN2 (n = 9) and CIN3 (n = 6). All the patients analyzed had at least one high-risk HPV serotype. The most frequently identified serotypes were 31 and 33 (n = 18 of 30). 6 or more HPV serotypes were found in 18 patients. Of 15 patients with high-grade lesions, 8 had the association of serotypes 31 and 33, and in 6 an isolated serotype was identified (16 and 51). CONCLUSIONS: The most frequently identified serotypes were 31 and 33. Unfortunately, the nonavalent vaccine that protects against the most frequent serotypes of HPV is not available in Latin America.

Embarazo triple monocorial espontáneo. A propósito de un caso / Spontaneous monochorionic trigemellar pregnancy. A case report / Gravidez monocoriônica espontânea tripla. Relato de caso

Resumen: El embarazo triple monocorial espontáneo tiene una incidencia muy baja, no identificándose factores causales que lo expliquen. La corionicidad determina el resultado perinatal de estas gestaciones, siendo la frecuencia de malos resultados mayor a medida que aumenta el número de fetos con una única placenta. Se presenta un caso de embarazo triple monocorial espontáneo cuyo seguimiento fue realizado en el Centro Hospitalario Pereira Rossell con excelente resultado perinatal.

Summary: Spontaneous monochorionic trigemellar pregnancies have a very low incidence, and there is no cause that may explain them. The chorionicity will determine the perinatal result of this gestations, but the frequency of poor results is higher as the number of fetuses sharing the same placenta increases. We present a case of a spontaneous monochorionic trigemellar gestation whose follow-up was carried out in the Pereira Rossell Pediatric Center with an excellent perinatal result.

Resumo: A tríplice gestação monocorial espontânea tem incidência muito baixa, não sendo identificados os fatores causais que a explicam. A corionicidade determina o resultado perinatal dessas gestações, e a frequência de resultados ruins é maior à medida que aumenta o número de fetos com uma única placenta. Apresentamos um caso de gravidez tríplice monocoriônica espontânea cujo seguimento foi realizado no Centro Hospitalar Pereira Rossell com excelentes resultados perinatais.

Ega3 from the fungal pathogen Aspergillus fumigatus is an endo-α-1,4-galactosaminidase that disrupts microbial biofilms.

Aspergillus fumigatus is an opportunistic fungal pathogen that causes both chronic and acute invasive infections. Galactosaminogalactan (GAG) is an integral component of the A. fumigatus biofilm matrix and a key virulence factor. GAG is a heterogeneous linear α-1,4-linked exopolysaccharide of galactose and GalNAc that is partially deacetylated after secretion. A cluster of five co-expressed genes has been linked to GAG biosynthesis and modification. One gene in this cluster, ega3, is annotated as encoding a putative α-1,4-galactosaminidase belonging to glycoside hydrolase family 114 (GH114). Herein, we show that recombinant Ega3 is an active glycoside hydrolase that disrupts GAG-dependent A. fumigatus and Pel polysaccharide-dependent Pseudomonas aeruginosa biofilms at nanomolar concentrations. Using MS and functional assays, we demonstrate that Ega3 is an endo-acting α-1,4-galactosaminidase whose activity depends on the conserved acidic residues, Asp-189 and Glu-247. X-ray crystallographic structural analysis of the apo Ega3 and an Ega3-galactosamine complex, at 1.76 and 2.09 Å resolutions, revealed a modified (ß/α)8-fold with a deep electronegative cleft, which upon ligand binding is capped to form a tunnel. Our structural analysis coupled with in silico docking studies also uncovered the molecular determinants for galactosamine specificity and substrate binding at the -2 to +1 binding subsites. The findings in this study increase the structural and mechanistic understanding of the GH114 family, which has >600 members encoded by plant and opportunistic human pathogens, as well as in industrially used bacteria and fungi.

Could human bocavirus be a causative agent of parotitis in children? / ¿Puede ser el bocavirus humano un agente causal de parotiditis en niños?

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