RESUMO
INTRODUCTION: Argentina authorized COVID-19 vaccination for adolescents 12 years and older in August 2021, and then for children three years and older in October 2021. Children aged 6 months-2 years received a two-dose regimen beginning July 2022. OBJECTIVE: This study aims to analyze the impact of COVID-19 vaccination among children aged 0-17, considering vaccination status and mortality for the 2020-2022 period. METHODS: We conducted a population-level analysis examining all-cause mortality, COVID-19 cases, deaths, and vaccination records. We compared outcomes with child mortality for diseases for which vaccination is compulsory, before and after each vaccine rollout. RESULTS: A decrease in COVID-19-related deaths was observed in 2022 for pediatric age groups (3-11 and 12-17) with relatively higher vaccination coverage. However, no decrease was observed for the 0-2 year old age group, which had the longest delay in access to immunization and lowest vaccination coverage. When compared to unvaccinated populations in 2022, we observe an 8-15-fold reduction in cumulative death rates for pediatric populations vaccinated with 1 or more doses, and a 16-18-fold reduction for those vaccinated with 2 or more doses. Historical analysis shows that for diseases for which vaccination is now compulsory in many countries, pre-vaccine-rollout mortality was lower than COVID-19 deaths during 2020-2022. CONCLUSIONS AND RELEVANCE: SARS-CoV-2 immunization was associated with reduced COVID-19 deaths for children and adolescents in Argentina. Our findings suggest that greater efforts should be undertaken to ensure wider COVID-19 vaccine coverage in children and adolescents, especially infants.
Assuntos
Vacinas contra COVID-19 , COVID-19 , SARS-CoV-2 , Vacinação , Humanos , Argentina/epidemiologia , COVID-19/mortalidade , COVID-19/prevenção & controle , COVID-19/epidemiologia , Criança , Adolescente , Pré-Escolar , Lactente , Vacinas contra COVID-19/administração & dosagem , Vacinação/estatística & dados numéricos , Masculino , Feminino , SARS-CoV-2/imunologia , Cobertura Vacinal/estatística & dados numéricos , Recém-NascidoRESUMO
Classical scoring functions may exhibit low accuracy in determining ligand binding affinity for proteins. The availability of both protein-ligand structures and affinity data make it possible to develop machine-learning models focused on specific protein systems with superior predictive performance. Here, we report a new methodology named SAnDReS that combines AutoDock Vina 1.2 with 54 regression methods available in Scikit-Learn to calculate binding affinity based on protein-ligand structures. This approach allows exploration of the scoring function space. SAnDReS generates machine-learning models based on crystal, docked, and AlphaFold-generated structures. As a proof of concept, we examine the performance of SAnDReS-generated models in three case studies. For all three cases, our models outperformed classical scoring functions. Also, SAnDReS-generated models showed predictive performance close to or better than other machine-learning models such as KDEEP, CSM-lig, and ΔVinaRF20. SAnDReS 2.0 is available to download at https://github.com/azevedolab/sandres.
Assuntos
Aprendizado de Máquina , Proteínas , Proteínas/química , Proteínas/metabolismo , Ligantes , Software , Simulação de Acoplamento MolecularRESUMO
The immune regulator gene AIRE plays an essential role in the establishment of immune tolerance and the prevention of autoimmunity. This transcription factor plays a critical role in promoting self-tolerance in the thymus by regulating the expression of a large number of self-antigens that share the common feature of being tissue-restricted in their expression pattern in the periphery. Dysfunction of AIRE in humans causes a rare disease, autoimmune polyglandular syndrome type 1 (APS1), characterized by an autoimmune response against peripheral tissues, particularly endocrine tissues. Although a few dominant mutations have been described, the inactivation of AIRE is usually caused by recessive mutations. Recent data suggests that alterations in AIRE function contribute not only to APS1 but also to more common forms of autoimmune disease. Here, we present a previously unreported missense mutation (NM_000383.2:c.260 T > C) in exon 2 of the AIRE gene, predicted to cause the substitution (p.(Leu87Pro)) in the CARD domain of the AIRE protein. When inherited in conjunction with another dysfunctional AIRE allele, this mutation was associated with immune dysregulation in a pediatric patient. The presence of hypergammaglobulinemia, malabsorption syndrome, ectodermal dysplasia, mucocutaneous candidiasis, vitiligo, and hypothyroidism as well as the presence of multiple autoantibodies allowed us to confirm an APS1 diagnosis.
Assuntos
Mutação de Sentido Incorreto , Poliendocrinopatias Autoimunes , Criança , Humanos , Proteína AIRE , Mutação , Poliendocrinopatias Autoimunes/genética , Poliendocrinopatias Autoimunes/diagnóstico , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
INTRODUÇÃO: Em geral, pacientes (p) incluídos em estudos randomizados são selecionados e exibem perfil clínico/angiográfico de menor gravidade do que os do mundo real. Na última década, ensaios clínicos envolvendo intervenções coronárias percutâneas (ICP) com novos modelos de stents farmacológicos (SF) tem primado por estabelecer apenas critérios clínicos de avaliação dos resultados, evitando a necessidade de reestudo angiográfico compulsório na evolução, o que pode atenuar discrepâncias significativas entre casos selecionados ou não para participar dos estudos, assim como permitir a inclusão de mais p. OBJETIVO: O objetivo desta investigação foi avaliar a hipótese supracitada. MÉTODOS: Recentemente foi realizada em nosso Serviço a inclusão de p diabéticos (PD) de forma prospectiva e randomizada num estudo que avaliou um novo modelo de SF, quando comparado a outro modelo utilizado na rotina. Havia critérios de exclusão restritos. Decidimos comparar os perfis clínico e angiográfico de PD incluídos ou não no estudo, finalizando aleatoriamente a amostra quando fosse incluído o centésimo p. Desta forma, entre 10/2020 e 07/2021, foram realizadas 336 ICP em PD de forma prospectiva, subdivididos em 2 grupos: A) 100(29%) PD incluídos no estudo; B) os 236(71%) restantes. Estabeleceram-se como significativos valores de p<0.05. RESULTADOS: Sexo, idade, diabetes dependente de insulina e extensão da doença coronária não diferiram entre os grupos. Predominaram significativamente no grupo B: disfunção renal crônica(21% vs 7%; p=0.0033), idade superior a 70 anos(37% vs 23% p=0.015), antecedente de ICP(32% vs 13%; p=<0.001), ICP para pontes de veia safena(5% vs 0% p=0.03) e infarto com elevação de ST(19% vs 8%; p=0.017); antecedente de cirurgia de revascularização miocárdica previa(10% vs 4% P=0.08) e ICP uniarterial (69% vs 57% p=0.054) também foram mais observados em B, mas não de forma significante. Predominaram no grupo A: ICP de múltiplos vasos (43% vs 25%; p=0.0016), apresentação clínica de angina estável (47% vs 31%; p=0.009) e revascularização completa (62% vs 47%; p=0.0016). Embora a permanência de UTI do grupo B foi maior (1.1(±3.5) vs 0.27(±1.5) P=0.024) os resultados imediatos da fase hospitalar, em conjunto ou individualmente, não diferiram. CONCLUSÕES: 1) Cerca de um em cada três PD tratados foram incluídos no estudo; 2) mesmo com critérios de inclusão/exclusão menos rígidos, observamos perfil clínico/angiográfico de menor complexidade nos casos incluídos.
Assuntos
Ensaios Clínicos Controlados Aleatórios como Assunto , Diabetes MellitusRESUMO
BACKGROUND: The idea of scoring function space established a systems-level approach to address the development of models to predict the affinity of drug molecules by those interested in drug discovery. OBJECTIVE: Our goal here is to review the concept of scoring function space and how to explore it to develop machine learning models to address protein-ligand binding affinity. METHOD: We searched the articles available in PubMed related to the scoring function space. We also utilized crystallographic structures found in the protein data bank (PDB) to represent the protein space. RESULTS: The application of systems-level approaches to address receptor-drug interactions allows us to have a holistic view of the process of drug discovery. The scoring function space adds flexibility to the process since it makes it possible to see drug discovery as a relationship involving mathematical spaces. CONCLUSION: The application of the concept of scoring function space has provided us with an integrated view of drug discovery methods. This concept is useful during drug discovery, where we see the process as a computational search of the scoring function space to find an adequate model to predict receptor-drug binding affinity.
RESUMO
A plethora of measures are being combined in the attempt to reduce SARS-CoV-2 spread. Due to its sustainability, contact tracing is one of the most frequently applied interventions worldwide, albeit with mixed results. We evaluate the performance of digital contact tracing for different infection detection rates and response time delays. We also introduce and analyze a novel strategy we call contact prevention, which emits high exposure warnings to smartphone users according to Bluetooth-based contact counting. We model the effect of both strategies on transmission dynamics in SERIA, an agent-based simulation platform that implements population-dependent statistical distributions. Results show that contact prevention remains effective in scenarios with high diagnostic/response time delays and low infection detection rates, which greatly impair the effect of traditional contact tracing strategies. Contact prevention could play a significant role in pandemic mitigation, especially in developing countries where diagnostic and tracing capabilities are inadequate. Contact prevention could thus sustainably reduce the propagation of respiratory viruses while relying on available technology, respecting data privacy, and most importantly, promoting community-based awareness and social responsibility. Depending on infection detection and app adoption rates, applying a combination of digital contact tracing and contact prevention could reduce pandemic-related mortality by 20-56%.
Assuntos
COVID-19/prevenção & controle , Busca de Comunicante/métodos , Smartphone , Humanos , Pandemias/prevenção & controle , SARS-CoV-2/patogenicidadeRESUMO
La presente carta al editor expresa una opinión novedosa sobre las perspectivas sobre la educación me´dica de pregrado durante la pandemia del COVID-19. Hace un repaso sobre la educación médica tradicional y sobre los beneficios y perjuicios de los nuevos modelos educacionales, como la telemedicina.
This letter expresses a novel opinion on the prospects for undergraduate medical education during the COVID-19 pandemic. It reviews traditional medical education and the benefits and harms of new educational models, such as telemedicine.
RESUMO
Experimental findings show the ubiquitous presence of graded responses and tuning curves in the neocortex, particularly in visual areas [1-15]. Among these, inferotemporal-cortex (IT) neurons respond to complex visual stimuli, but differences in the neurons' responses can be used to distinguish the stimuli eliciting the responses [8, 9, 16-18]. The IT projects directly to the medial temporal lobe (MTL) [19], where neurons respond selectively to different pictures of specific persons and even to their written and spoken names [20-22]. However, it is not clear whether this is done through a graded coding, as in the neocortex, or a truly invariant code, in which the response-eliciting stimuli cannot be distinguished from each other. To address this issue, we recorded single neurons during the repeated presentation of different stimuli (pictures and written and spoken names) corresponding to the same persons. Using statistical tests and a decoding approach, we found that only in a minority of cases can the different pictures of a given person be distinguished from the neurons' responses and that in a larger proportion of cases, the responses to the pictures were different to the ones to the written and spoken names. We argue that MTL neurons tend to lack a representation of sensory features (particularly within a sensory modality), which can be advantageous for the memory function attributed to this area [23-25], and that a full representation of memories is given by a combination of mostly invariant coding in the MTL with a representation of sensory features in the neocortex.
Assuntos
Hipocampo/fisiologia , Neocórtex/fisiologia , Neurônios/fisiologia , Reconhecimento Visual de Modelos/fisiologia , Lobo Temporal/fisiologia , Adulto , Argentina , Mapeamento Encefálico , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Célula Única , Adulto JovemRESUMO
DHHC palmitoyltransferases (DHHC-PATs) are very peculiar in that, outside the DHHC domain, they are very divergent even across orthologs from closely related species. This represents a challenge for the bioinformatic analyses of these proteins. Sequence-based analyses and predictions require a valid sequence alignment, which for this family of proteins requires extensive manual curation and this is difficult to attain for the nonspecialist. Here we present a simple method for the in silico analysis of the sequence of a particular PAT, that would allow for the identification of important structural features and functional residues in a PAT or PAT family.
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Acetiltransferases , Alinhamento de Sequência , Análise de Sequência de Proteína , Software , Acetiltransferases/química , Acetiltransferases/genética , Motivos de Aminoácidos , Biologia ComputacionalRESUMO
RESUMEN: Objetivo: Se desarrolló una investigación que tuvo por propósito determinar el patrón de reabsorción del tejido óseo de implantes rehabilitados con coronas de acrílico que actúan como pilares de una prótesis parcial removible con retenedores extracoronarios. Materiales y métodos: Para medir tal efecto se diseñó un dispositivo que permitió estandarizar el registro del nivel óseo mediante una radiografía retroalveolar con técnica de paralelismo, considerando que este procedimiento permite observar gran cantidad de detalles, ofreciendo una imagen de alta definición, con una mínima distorsión, bajo costo y mínimo riesgo de radiación. Los resultados: indican que en un intervalo de seis meses medido después de realizada la rehabilitación a los 24 y 30 meses respectivamente se observa una reabsorción ósea que en promedio alcanza 0,12 mm siendo esta diferencia estadísticamente significativa entre ambos tiempos de registros. Conclusión: Los patrones de reabsorción de nuestra investigación son similares a los que ocurren con otras formas de rehabilitación sobre implantes oseointegrados.
ABSTRACT: Objetive: The purpose of this research was to determine the pattern of reabsorption of bone tissue from rehabilitated implants with acrylic crowns that act as supports of a removable partial denture with extra-coronary retainers. Material and method: To measure this effect, a device was designed to standardize the measurement of the bone level by a retro-alveolar radiography with parallelism exposure technique, considering that this procedure allows to observe great amount of details, offering an image of high definition, with a minimum distortion, low cost and minimal risk of radiation. The results: indicate that in a six-month interval measured after rehabilitation at 24 and 30 months respectively a bone resorption is observed, which on average reaches 0.12 mm, this difference being statistically significant between both recording times. Conclusion: The resorption patterns of our research are similar to those that occur with other forms of rehabilitation on osseointegrated implants.
Assuntos
Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Idoso , Implantes Dentários , Boca Edêntula , Coroas , Prótese Parcial Removível , Mandíbula , Estudos LongitudinaisRESUMO
Autodock Vina is a very popular, and highly cited, open source docking program. Here we present a scoring function which we call Vinardo (Vina RaDii Optimized). Vinardo is based on Vina, and was trained through a novel approach, on state of the art datasets. We show that the traditional approach to train empirical scoring functions, using linear regression to optimize the correlation of predicted and experimental binding affinities, does not result in a function with optimal docking capabilities. On the other hand, a combination of scoring, minimization, and re-docking on carefully curated training datasets allowed us to develop a simplified scoring function with optimum docking performance. This article provides an overview of the development of the Vinardo scoring function, highlights its differences with Vina, and compares the performance of the two scoring functions in scoring, docking and virtual screening applications. Vinardo outperforms Vina in all tests performed, for all datasets analyzed. The Vinardo scoring function is available as an option within Smina, a fork of Vina, which is freely available under the GNU Public License v2.0 from http://smina.sf.net. Precompiled binaries, source code, documentation and a tutorial for using Smina to run the Vinardo scoring function are available at the same address.
Assuntos
Avaliação Pré-Clínica de Medicamentos , Simulação de Acoplamento Molecular , Software , Bases de Dados como Assunto , Ligantes , TermodinâmicaRESUMO
It is still unclear why some proteins that travel along the secretory pathway are retained in the Golgi complex whereas others make their way to the plasma membrane. Recent bioinformatic analyses on a large number of single-spanning membrane proteins support the hypothesis that specific features of the transmembrane domain (TMD) are relevant to the sorting of these proteins to particular organelles. Here we experimentally test this hypothesis for Golgi and plasma membrane proteins. Using the Golgi SNARE protein Sft1 and the plasma membrane SNARE protein Sso1 from Saccharomyces cerevisiae as model proteins, we modified the length of their TMDs and the volume of their exoplasmic hemi-TMD, and determined their subcellular localization both in yeast and mammalian cells. We found that short TMDs with high-volume exoplasmic hemi-TMDs confer Golgi membrane residence, whereas TMDs with low-volume exoplasmic hemi-TMDs, either short or long, confer plasma membrane residence to these proteins. Results indicate that the shape of the exoplasmic hemi-TMD, in addition to the length of the entire TMD, determine retention in the Golgi or exit to the plasma membrane of Type II membrane proteins.
Assuntos
Regulação Fúngica da Expressão Gênica , Complexo de Golgi/metabolismo , Proteínas de Membrana/química , Proteínas Qa-SNARE/química , Proteínas Qc-SNARE/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/metabolismo , Sequência de Aminoácidos , Animais , Células CHO , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Cricetulus , Complexo de Golgi/ultraestrutura , Proteínas de Membrana/metabolismo , Dados de Sequência Molecular , Ligação Proteica , Estrutura Terciária de Proteína , Transporte Proteico , Proteínas Qa-SNARE/genética , Proteínas Qa-SNARE/metabolismo , Proteínas Qc-SNARE/genética , Proteínas Qc-SNARE/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismoRESUMO
S-acylation, commonly known as palmitoylation, is a widespread post-translational modification of proteins that consists of the thioesterification of one or more cysteine residues with fatty acids. This modification is catalysed by a family of PATs (palmitoyltransferases), characterized by the presence of a 50-residue long DHHC-CRD (Asp-His-His-Cys cysteine-rich domain). To gain knowledge on the structure-function relationships of these proteins, we carried out a random-mutagenesis assay designed to uncover essential amino acids in Swf1, the yeast PAT responsible for the palmitoylation of SNARE (soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor) proteins. We identified 21 novel loss-of-function mutations, which are mostly localized within the DHHC-CRD. Modelling of the tertiary structure of the Swf1 DHHC domain suggests that it could fold as a zinc-finger domain, co-ordinating two zinc atoms in a CCHC arrangement. All residues predicted to be involved in the co-ordination of zinc were found to be essential for Swf1 function in the screen. Moreover, these mutations result in unstable proteins, in agreement with a structural role for these zinc fingers. The conservation of amino acids predicted to form each zinc-binding pocket suggests a shared function, as the selective pressure to maintain them is lost upon mutation of one of them. A Swf1 orthologue that lacks one of the zinc-binding pockets is able to complement a yeast swf1∆ strain, possibly because a similar fold can be stabilized by hydrogen bonds instead of zinc co-ordination. Finally, we show directly that recombinant Swf1 DHHC-CRD is able to bind zinc. Sequence analyses of DHHC domains allowed us to present models of the zinc-binding properties for all PATs.
Assuntos
Aciltransferases/química , Proteínas Fúngicas/química , Yarrowia/enzimologia , Zinco/química , Aciltransferases/genética , Aciltransferases/metabolismo , Substituição de Aminoácidos , Sítios de Ligação , Complexos de Coordenação/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Técnicas de Inativação de Genes , Teste de Complementação Genética , Humanos , Lipoilação , Filogenia , Ligação Proteica , Estrutura Terciária de Proteína , Saccharomyces cerevisiae , Análise de Sequência de Proteína , Homologia Estrutural de ProteínaRESUMO
BACKGROUND: The aim of this study was to document the types of diseases treated by the use of medicinal plants, their main applications and also to have a report of the major diseases treated at the Hospital of San Pablo de Huacareta (Chuquisaca Bolivia). METHODS: We conducted semi-structured interviews on the use medicinal plants with 10 local informants, and categorized the kinds of diseases treated by traditional medicine. We obtained reports of cases treated at the Hospital of Huacareta in order to compare the use frequency of traditional medicine and allopathic medicine for the treatment of recurrent diseases in the area. RESULTS: Our survey identified 258 traditional medicine uses, spanning a total of 13 diseases categories and including 91 native and exotic plant species and one unidentified sample plant type. Gastrointestinal disorders (55%) were most frequently treated with medicinal plants, followed by afflictions of the musculoskeletal system (25%) and dermatological disorders (24%). Hospital information indicates that the most common diseases are acute respiratory infections (47%) and acute diarrheal diseases (37%). The herbal remedies were mostly used in the form of teas and decoctions. The informants used mainly native plant species, although exotic species has been introduced to the pharmacopoeia. CONCLUSIONS: The treatment of gastrointestinal disorders is the primary objective of the medical ethnobotany of the inhabitants of Huacareta, while respiratory system diseases are mostly treated in the hospital. Looking at the data from the Hospital records we can infer that gastrointestinal disorders are among the most common diseases in the study area. For most respondents, traditional medicine is a reliable choice for the care of their illnesses. However, the preference of the population for either traditional medicine or allopathic medicine needs to be clarified in future comparative studies to obtain more convincing results. The results presented can be used as a base for subsequent work related to traditional medicine and its contribution to allopathic medicine in San Pablo de Huacareta.
Assuntos
Etnobotânica , Gastroenteropatias/tratamento farmacológico , Hospitais , Medicina Tradicional , Fitoterapia , Preparações de Plantas/uso terapêutico , Doenças Respiratórias/tratamento farmacológico , Idoso , Bolívia , Diarreia/tratamento farmacológico , Feminino , Humanos , Entrevistas como Assunto , Masculino , Pessoa de Meia-Idade , Doenças Musculoesqueléticas/tratamento farmacológico , Aceitação pelo Paciente de Cuidados de Saúde , Plantas Medicinais , Dermatopatias/tratamento farmacológicoRESUMO
In Giardia, lysosome-like peripheral vacuoles (PVs) need to specifically coordinate their endosomal and lysosomal functions to be able to successfully perform endocytosis, protein degradation and protein delivery, but how cargo, ligands and molecular components generate specific routes to the PVs remains poorly understood. Recently, we found that delivering membrane Cathepsin C and the soluble acid phosphatase (AcPh) to the PVs is adaptin (AP1)-dependent. However, the receptor that links AcPh and AP1 was never described. We have studied protein-binding to AcPh by using H6-tagged AcPh, and found that a membrane protein interacted with AcPh. This protein, named GlVps (for Giardia lamblia Vacuolar protein sorting), mainly localized to the ER-nuclear envelope and in some PVs, probably functioning as the sorting receptor for AcPh. The tyrosine-binding motif found in the C-terminal cytoplasmic tail domain of GlVps was essential for its exit from the endoplasmic reticulum and transport to the vacuoles, with this motif being necessary for the interaction with the medium subunit of AP1. Thus, the mechanism by which soluble proteins, such as AcPh, reach the peripheral vacuoles in Giardia appears to be very similar to the mechanism of lysosomal protein-sorting in more evolved eukaryotic cells.
Assuntos
Giardia lamblia/metabolismo , Vacúolos/metabolismo , Fosfatase Ácida/metabolismo , Animais , Catepsina C/metabolismoRESUMO
We report a study of concurrent eye movements and electroencephalographic (EEG) recordings while subjects freely explored a search array looking for hidden targets. We describe a sequence of fixation-event related potentials (fERPs) that unfolds during â¼ 400 ms following each fixation. This sequence highly resembles the event-related responses in a replay experiment, in which subjects kept fixation while a sequence of images occurred around the fovea simulating the spatial and temporal patterns during the free viewing experiment. Similar responses were also observed in a second control experiment where the appearance of stimuli was controlled by the experimenters and presented at the center of the screen. We also observed a relatively early component (â¼150 ms) that distinguished between targets and distractors only in the freeviewing condition. We present a novel approach to match the critical properties of two conditions (targets/distractors), which can be readily adapted to other paradigms to investigate EEG components during free eye-movements.
Assuntos
Eletroencefalografia/métodos , Potenciais Evocados Visuais/fisiologia , Movimentos Oculares/fisiologia , Fixação Ocular/fisiologia , Percepção de Forma/fisiologia , Adulto , Atenção/fisiologia , Feminino , Fóvea Central/fisiologia , Humanos , Masculino , Estimulação Luminosa/métodos , Tempo de Reação/fisiologia , Percepção Espacial/fisiologia , Adulto JovemRESUMO
Many proteins are modified after their synthesis, by the addition of a lipid molecule to one or more cysteine residues, through a thioester bond. This modification is called S-acylation, and more commonly palmitoylation. This reaction is carried out by a family of enzymes, called palmitoyltransferases (PATs), characterized by the presence of a conserved 50- aminoacids domain called "Asp-His-His-Cys- Cysteine Rich Domain" (DHHC-CRD). There are 7 members of this family in the yeast Saccharomyces cerevisiae, and each of these proteins is thought to be responsible for the palmitoylation of a subset of substrates. Substrate specificity of PATs, however, is not yet fully understood. Several yeast PATs seem to have overlapping specificity, and it has been proposed that the machinery responsible for palmitoylating peripheral membrane proteins in mammalian cells, lacks specificity altogether.Here we investigate the specificity of transmembrane protein palmitoylation in S. cerevisiae, which is carried out predominantly by two PATs, Swf1 and Pfa4. We show that palmitoylation of transmembrane substrates requires dedicated PATs, since other yeast PATs are mostly unable to perform Swf1 or Pfa4 functions, even when overexpressed. Furthermore, we find that Swf1 is highly specific for its substrates, as it is unable to substitute for other PATs. To identify where Swf1 specificity lies, we carried out a bioinformatics survey to identify amino acids responsible for the determination of specificity or Specificity Determination Positions (SDPs) and showed experimentally, that mutation of the two best SDP candidates, A145 and K148, results in complete and partial loss of function, respectively. These residues are located within the conserved catalytic DHHC domain suggesting that it could also be involved in the determination of specificity. Finally, we show that modifying the position of the cysteines in Tlg1, a Swf1 substrate, results in lack of palmitoylation, as expected for a highly specific enzymatic reaction.
Assuntos
Acetiltransferases/metabolismo , Lipoilação/fisiologia , Proteínas de Membrana/metabolismo , Leveduras/metabolismo , Acetiltransferases/química , Acetiltransferases/genética , Acetiltransferases/fisiologia , Aciltransferases/química , Aciltransferases/genética , Aciltransferases/metabolismo , Aciltransferases/fisiologia , Sequência de Aminoácidos , Domínio Catalítico/genética , Domínio Catalítico/fisiologia , Lipoilação/genética , Proteínas de Membrana/química , Modelos Biológicos , Dados de Sequência Molecular , Estrutura Terciária de Proteína/fisiologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Recombinantes de Fusão/fisiologia , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologia , Homologia de Sequência de Aminoácidos , Especificidade por Substrato/genética , Leveduras/genéticaRESUMO
Glycolipids constitute a complex family of amphipathic molecules structurally characterized by a hydrophilic mono- or oligo-saccharide moiety linked to a hydrophobic ceramide moiety. Due to their asymmetric distribution in cell membranes, exposing the saccharide moiety to the extracytoplasmic side of the cell, glycolipids participate in a variety of cell-cell and cell-ligand interactions. Here we summarize aspects of the cell biology of the stepwise synthesis of the saccharide moiety in the Golgi complex of cells from vertebrates. In particular we refer to the participant glycosyltransferases, with emphasis on their trafficking along the secretory pathway, their retention and organization in the Golgi complex membranes and their dependence on the Golgi complex ultra structural organization for proper function.
Assuntos
Glicolipídeos/química , Complexo de Golgi/metabolismo , Oligossacarídeos/biossíntese , Animais , Retículo Endoplasmático/metabolismo , Glicosiltransferases/química , Glicosiltransferases/metabolismo , Complexo de Golgi/enzimologia , HumanosRESUMO
Brain tissue is characterized by its high glycosphingolipid content, particularly those containing sialic acid (gangliosides). As a result of this observation, brain tissue was a focus for studies leading to the characterization of the enzymes participating in ganglioside biosynthesis, and their participation in driving the compositional changes that occur in glycolipid expression during brain development. Later on, this focus shifted to the study of cellular aspects of the synthesis, which lead to the identification of the site of synthesis in the neuronal soma and their axonal transport toward the periphery. In this review article, we will focus in subcellular aspects of the biosynthesis of glycosphingolipid oligosaccharides, particularly the mechanisms underlying the trafficking of glycosphingolipid glycosyltransferases from the endoplasmic reticulum to the Golgi, those that promote their retention in the Golgi and those that participate in their topological organization as part of the complex membrane bound machinery for the synthesis of glycosphingolipids.