Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 7 de 7
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
mSystems ; 5(6)2020 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-33172970

RESUMO

Marine microbes are known to degrade hydrocarbons; however, microbes inhabiting deep-sea sediments remain largely unexplored. Previous studies into the classical pathways of marine microbial metabolism reveal diverse chemistries; however, metabolic profiling of marine microbes cultured with hydrocarbons is limited. In this study, taxonomic (amplicon sequencing) profiles of two environmental deep-sea sediments (>1,200 m deep) were obtained, along with taxonomic and metabolomic (mass spectrometry-based metabolomics) profiles of microbes harbored in deep-sea sediments cultured with hydrocarbons as the sole energy source. Samples were collected from the Gulf of México (GM) and cultured for 28 days using simple (toluene, benzene, hexadecane, and naphthalene) and complex (petroleum API 40) hydrocarbon mixtures as the sole energy sources. The sediment samples harbored diverse microbial communities predominantly classified into Woeseiaceae and Kiloniellaceae families, whereas Pseudomonadaceae and Enterobacteriaceae families prevailed after sediments were cultured with hydrocarbons. Chemical profiling of microbial metabolomes revealed diverse chemical groups belonging primarily to the lipids and lipid-like molecules superclass, as well as the organoheterocyclic compound superclass (ClassyFire annotation). Metabolomic data and prediction of functional profiles indicated an increase in aromatic and alkane degradation in samples cultured with hydrocarbons. Previously unreported metabolites, identified as intermediates in the degradation of hydrocarbons, were annotated as hydroxylated polyunsaturated fatty acids and carboxylated benzene derivatives. In summary, this study used mass spectrometry-based metabolomics coupled to chemoinformatics to demonstrate how microbes from deep-sea sediments could be cultured in the presence of hydrocarbons. This study also highlights how this experimental approach can be used to increase the understanding of hydrocarbon degradation by deep-sea sediment microbes.IMPORTANCE High-throughput technologies and emerging informatics tools have significantly advanced knowledge of hydrocarbon metabolism by marine microbes. However, research into microbes inhabiting deep-sea sediments (>1,000 m) is limited compared to those found in shallow waters. In this study, a nontargeted and nonclassical approach was used to examine the diversity of bacterial taxa and the metabolic profiles of hydrocarbon-degrading deep-sea microbes. In conclusion, this study used metabolomics and chemoinformatics to demonstrate that microbes from deep-sea sediment origin thrive in the presence of toxic and difficult-to-metabolize hydrocarbons. Notably, this study provides evidence of previously unreported metabolites and the global chemical repertoire associated with the metabolism of hydrocarbons by deep-sea microbes.

2.
J Proteomics ; 218: 103689, 2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32088355

RESUMO

Hemocyanin (Hc) is a multifunctional macromolecule involved in oxygen transport and non-specific immunity in shrimp. Hc is crucial in physiology and nutrition linked with optimal performance in aquaculture production systems. In medicine, Hc has been approved for clinical use in humans as adjuvant and anticancer therapeutic. In contrast, Hc has also been identified as one of the proteins causing anaphylaxis following shrimp consumption. The role of individual Hc isoforms remains unknown due to a lack of resolved Hc isoforms. We successfully identified eleven different Penaeus monodon hemocyanin (PmoHc) γ isoforms including two truncated isoforms (50 and 20 kDa) and one PmoHc ß isoform in haemolymph using proteomics informed by transcriptomics. Amino acid sequence homology ranged from 24 to 97% between putative PmoHc gene isoforms. Hc isoforms showed specific patterns of transcript expression in shrimp larval stages and adult hepatopancreas. These findings enable isoform level investigations aiming to define molecular mechanisms underpinning Hc functionality in shrimp physiology and immunity, as well as their individual immunogenic role in human allergy. Our research demonstrates the power of proteomics informed by transcriptomics to resolve isoform complexity in non-model organisms and lay the foundations for improved performance within the aquaculture industry and advance allergenic applications in medicine. SIGNIFICANCE: The roles of hemocyanin (Hc) in shrimp homeostasis and immunity as well as in human allergy are not well understood because the complexity of Hc isoforms has remained unresolved. Our results have confirmed the existence of at least 12 individual Hc isoforms in shrimp haemolymph and validated putative Hc gene assemblies from transcriptomics. Our findings will enable monitoring the expression of specific Hc isoforms in shrimp haemolymph during different environmental, nutritional and pathogenic conditions, thus providing insights into isoform specific functional roles. In medicine, the potential allergenicity of each Hc isoform could be determined and advance allergenic applications. Lastly, since Hc comprises up to 95% of the total protein in haemolymph, these isoforms become ideal targets for prawn provenance, traceability and food contamination studies.

3.
Rapid Commun Mass Spectrom ; 34(9): e8723, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31922636

RESUMO

RATIONALE: Cytokines are cell regulatory molecules of high importance as indicators for homeostasis and pathology in many species. The current method to measure cytokines in body fluids is reagent dependent, requiring highly specific paired antibodies. METHODS: A liquid chromatography/multiple reaction monitoring mass spectrometry (LC/MRM-MS)-based approach was developed to simultaneously establish the limits of detection (LODs) and quantification (LOQs) for recombinant cytokines IL-1ß, IL-6, IFNγ and TNFα as pure standards and in bovine sera. All experimental LC/MRM-MS data are available at CSIRO Data Access Portal repository under identifier doi.org/10.25919/5de8a0232a862. RESULTS: The present method enabled LODs and LOQs as low as 1.05 and 1.12 fmol/µL in the experiment comprised of pure standards. Comparable results were obtained in the experiment where digested cytokines were mixed with pre-digested sera proteins. The intrinsic matrix effects were evident when intact cytokines were co-digested within undiluted and undigested sera decreasing the ability to detect and quantify cytokines by 10,000-fold compared with pure standards and pre-digested sera. CONCLUSIONS: The developed LC/MRM-MS method provided insights into the difficulties in detecting the target peptides when embedded in complex matrices. Nonetheless, the method may potentially be readily applied in biomarker-focused research interrogating fluids of lesser complexity such as synovial fluid, cerebrospinal fluid and tissue culture media.

4.
J Proteome Res ; 18(9): 3342-3352, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31321981

RESUMO

The freshwater snail Pomacea canaliculata, an invasive species of global significance, possesses a well-developed digestive system and diverse feeding mechanisms enabling the intake of a wide variety of food. The identification of glycosidases in adult snails would increase the understanding of their digestive physiology and potentially generate new opportunities to eradicate and/or control this invasive species. In this study, liquid chromatography coupled to tandem mass spectrometry was applied to define the occurrence, diversity, and origin of glycoside hydrolases along the digestive tract of P. canaliculata. A range of cellulases, hemicellulases, amylases, maltases, fucosidases, and galactosidases were identified across the digestive tract. The digestive gland and the contents of the crop and style sac yield a higher diversity of glycosidase-derived peptides. Subsequently, peptides derived from 81 glycosidases (46 proteins from the public database and 35 uniquely from the transcriptome database) that were distributed among 13 glycoside hydrolase families were selected and quantified using multiple reaction monitoring mass spectrometry. This study showed a high glycosidase abundance and diversity in the gut contents of P. canaliculata which participate in extracellular digestion of complex dietary carbohydrates. Salivary and digestive glands were the main tissues involved in their synthesis and secretion.


Assuntos
Glicosídeo Hidrolases/genética , Proteômica , Caramujos/genética , Transcriptoma/genética , Animais , Cromatografia Líquida/métodos , Trato Gastrointestinal/metabolismo , Glicosídeo Hidrolases/isolamento & purificação , Glicosídeo Hidrolases/metabolismo , Espécies Introduzidas , Caramujos/metabolismo , Espectrometria de Massas em Tandem/métodos
5.
Food Chem ; 254: 302-308, 2018 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-29548457

RESUMO

A strict, lifelong gluten-free (GF) diet is currently the only treatment for coeliac disease (CD). Vinegar and soy sauce are fermented condiments that often include wheat and/or barley. During fermentation cereal proteins are partially degraded by enzymes to yield peptide fragments and amino acids. Whether these fermented products contain intact or degraded gluten proteins and if they are safe for people with CD remains in question. LC-MS offers the benefit of being able to detect hydrolysed gluten that might be present in commercial vinegar and soy sauce products. LC-MS revealed the presence of gluten in malt vinegar, wherein the identified peptides derived from B-, D- and γ-hordein from barley, as well as γ-gliadin, and HMW- and LMW-glutenins from wheat that are known to contain immunopathogenic epitopes. No gluten was detected in the soy sauces examined despite wheat being a labelled ingredient indicating extensive hydrolysis of gluten during soy sauce production.


Assuntos
Ácido Acético/análise , Alimentos e Bebidas Fermentados/análise , Glutens/análise , Alimentos de Soja/análise , Espectrometria de Massas em Tandem/métodos , Cromatografia Líquida , Análise de Alimentos/métodos , Gliadina/análise , Hordeum/química , Hidrólise , Peso Molecular , Peptídeos/análise , Triticum/química
6.
Artigo em Inglês | MEDLINE | ID: mdl-27268288

RESUMO

Inefficient control of temperate abalone spawning prevents pair-wise breeding and production of abalone with highly marketable traits. Traditionally, abalone farmers have used a combination of UV irradiation and application of temperature gradients to the tank water to artificially induce spawning. Proteins are known to regulate crucial processes such as respiration, muscle contraction, feeding, growth and reproduction. Spawning as a pre-requisite of abalone reproduction is likely to be regulated, in part, by endogenous proteins. A first step in elucidating the mechanisms that regulate spawning is to identify which proteins are directly involved during spawning. The present study examined protein expression following traditional spawning induction in the Haliotis laevigata female. Gonads were collected from abalone in the following physiological states: (1) spawning; (2) post-spawning; and (3) failed-to-spawn. Differential protein abundance was initially assessed using two-dimensional difference in-gel electrophoresis coupled with mass spectrometry for protein identification. A number of reproductive proteins such as vitellogenin, vitelline envelope zona pellucida domain 29 and prohibitin, and metabolic proteins such as thioredoxin peroxidase, superoxide dismutase and heat shock proteins were identified. Differences in protein abundance levels between physiological states were further assessed using scheduled multiple reaction monitoring mass spectrometry. Positive associations were observed between the abundance of specific proteins, such as heat shock cognate 70 and peroxiredoxin 6, and the propensity or failure to spawn in abalone. These findings have contributed to better understand both the effects of oxidative and heat stress over abalone physiology and their influence on abalone spawning.


Assuntos
Gastrópodes/genética , Gastrópodes/metabolismo , Gônadas/metabolismo , Proteoma/análise , Reprodução/fisiologia , Animais , Aquicultura , Eletroforese em Gel Bidimensional , Feminino , Gastrópodes/fisiologia , Perfilação da Expressão Gênica , Gônadas/química , Proteoma/genética , Proteoma/metabolismo , Reprodução/genética
7.
J Proteomics ; 108: 337-53, 2014 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-24929219

RESUMO

UNLABELLED: Aside from their critical role in reproduction, abalone gonads serve as an indicator of sexual maturity and energy balance, two key considerations for effective abalone culture. Temperate abalone farmers face issues with tank restocking with highly marketable abalone owing to inefficient spawning induction methods. The identification of key proteins in sexually mature abalone will serve as the foundation for a greater understanding of reproductive biology. Addressing this knowledge gap is the first step towards improving abalone aquaculture methods. Proteomic profiling of female and male gonads of greenlip abalone, Haliotis laevigata, was undertaken using liquid chromatography-mass spectrometry. Owing to the incomplete nature of abalone protein databases, in addition to searching against two publicly available databases, a custom database comprising genomic data was used. Overall, 162 and 110 proteins were identified in females and males respectively with 40 proteins common to both sexes. For proteins involved in sexual maturation, sperm and egg structure, motility, acrosomal reaction and fertilization, 23 were identified only in females, 18 only in males and 6 were common. Gene ontology analysis revealed clear differences between the female and male protein profiles reflecting a higher rate of protein synthesis in the ovary and higher metabolic activity in the testis. BIOLOGICAL SIGNIFICANCE: A comprehensive mass spectrometry-based analysis was performed to profile the abalone gonad proteome providing the foundation for future studies of reproduction in abalone. Key proteins involved in both reproduction and energy balance were identified. Genomic resources were utilised to build a database of molluscan proteins yielding >60% more protein identifications than in a standard workflow employing public protein databases.


Assuntos
Bases de Dados de Proteínas , Gastrópodes/metabolismo , Ovário/metabolismo , Proteômica , Testículo/metabolismo , Animais , Feminino , Gastrópodes/genética , Masculino , Reprodução/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...