Q112431179
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The aim of this study was to compare the microbiological quality of cooked sausages produced with a traditional salt content (2.1%) and reformulated batches with a salt content reduced to 1.7%. The reformulation was tested on two types of comminuted meat products – Špekáčky sausage with a diameter of up to 46 mm or Bologna-type sausages in diameter of 85 mm (Gothaj sausage) or 75 mm (Junior sausage). The total viable count (TVC) increased only slightly during the four-week storage (4 ± 1 °C) of all batches of Špekáčky sausage. Comparing batches 1.7 and 2.1, there is an evident difference in the number of CFU/g, with samples of Špekáčky 1.7 showing numbers of bacteria higher by approximately 1 logarithmic order throughout practically the entire storage period (P = 0.001). The population of lactic acid bacteria (LAB) remained well beneath a value of 5.0 log CFU/g even at the end of the experiment. For Bologna-type sausages, the TVC was either beneath the limit of detection or at its boundary in all samples. LAB were not detected during storage of Bologna-type sausages. The results confirmed that the proportion of salt in cooked sausages can be reduced to 1.7% without negatively affecting the shelf life or safety of the final products.
Thirteen coagulase-negative, oxidase-negative, and novobiocin-susceptible staphylococci were isolated from human clinical specimens. The isolates were differentiated from known staphylococcal species on the basis of 16S rRNA, hsp60, rpoB, dnaJ, tuf, and gap gene sequencing, automated ribotyping, (GTG)5-PCR fingerprinting, and MALDI-TOF MS analysis. Phylogenetic analysis based on the 16S rRNA gene sequence indicated phylogenetic relatedness of the analyzed strains to Staphylococcus haemolyticus, Staphylococcus hominis, Staphylococcus devriesei, and Staphylococcus lugdunensis. DNA-DNA hybridization experiments between representative strains CCM 8418(T), CCM 8421(T), and the closest phylogenetic neighbors confirmed that the isolates represent novel Staphylococcus species, for which the name Staphylococcus petrasii sp. nov. is proposed. Genotypic and phenotypic analyses unambiguously split the strains into two closely related subclusters. Based on the results, two novel subspecies S. petrasii subsp. petrasii subsp. nov. and S. petrasii subsp. croceilyticus subsp. nov. are proposed, with type strains CCM 8418(T) (=CCUG 62727(T)) and CCM 8421(T) (=CCUG 62728(T)), respectively.
- MeSH
- antibakteriální látky metabolismus MeSH
- bakteriální proteiny genetika MeSH
- DNA bakterií chemie genetika MeSH
- fylogeneze MeSH
- hybridizace nukleových kyselin MeSH
- koagulasa metabolismus MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- novobiocin metabolismus MeSH
- oxidoreduktasy metabolismus MeSH
- ribozomální DNA chemie genetika MeSH
- RNA ribozomální 16S genetika MeSH
- sekvenční analýza DNA MeSH
- shluková analýza MeSH
- stafylokokové infekce mikrobiologie MeSH
- Staphylococcus klasifikace genetika izolace a purifikace metabolismus MeSH
- techniky typizace bakterií MeSH
- ucho mikrobiologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Biochemical characteristics of patatin proteins purified by ion-exchange and affinity chromatography from tubers of 20 potato cultivars were studied to evaluate their genotype differences with respect to utility groups, table potato cultivars (TPCs) and processing potato cultivars (PPCs). Both groups of cultivars showed similar values of protein content in dry matter (3.98-7.39%) and of patatin relative abundance (5.40-35.40%). Three mass levels (∼40.6, 41.8, and 42.9 kDa) of purified patatins were found by MALDI-TOF MS within all cultivars. Differences among mass levels corresponding with the mass of sugar antenna (∼1.2 kDa) confirmed the previous concept of different glycosylation extentsin patatin proteins. It was showed that the individual types of patatin varying in their masses occur in the patatin family in a ratio specific for each of the cultivars, with the lowest mass type being the major one. Electrophoretic analyses demonstrated wide cultivar variability in number of patatin forms. Especially 2D-PAGE showed 17-23 detected protein spots independently on the utility group. Specific lipid acyl hydrolase (LAH) activity of purified patatins from the individual tested cultivars varied between 0.92 and 5.46 μmol/(min mg). Patatin samples within most of the TPCs exhibited higher values of specific LAH activity than samples of PPCs. It may be supposed that individual patatin forms do not have similar physiological roles.
- MeSH
- druhová specificita MeSH
- glykosylace MeSH
- hlízy rostlin chemie MeSH
- karboxylesterhydrolasy chemie izolace a purifikace metabolismus MeSH
- molekulová hmotnost MeSH
- protein - isoformy chemie izolace a purifikace metabolismus MeSH
- rostlinné proteiny analýza chemie izolace a purifikace metabolismus MeSH
- Solanum tuberosum chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
Východiska: Termín biomarker se v posledních letech stal, zvláště pak ve spojení s termínem klinická proteomika, jedním z nejfrekventovanějších pojmů v oblasti biomedicínského výzkumu. Předmětem této práce byl výběr vhodné metody frakcionace krevní plazmy umožňující oddělení majoritních proteinů od nízkoabundantní proteinové frakce před následnou proteomickou analýzou pomocí dvourozměrné gelové elektroforézy (2-DE) s cílem zlepšit rozlišení 2-DE mapy a identifikaci proteinů hmotnostní spektrometrií. Materiál a metody: Nejprve bylo provedeno paralelní srovnání dvou metod frakcionace krevní plazmy (depleční kolona MARS vs ProteoMiner) na souboru 10 vzorků. Na základě výsledků srovnávacích experimentů byla k analýze vzorků 18 pacientů nemocných mnohočetným myelomem a léčených režimy s bortezomibem použita frakcionační metoda ProteoMiner v kombinaci s 2-DE. Soubor pacientů byl rozdělen na dvě skupiny: skupina pacientů rezistentních na chemoterapii (9 pacientů – progrese onemocnění, stabilizace onemocnění) a skupina pacientů s pozitivní léčebnou odpovědí (9 pacientů – kompletní a parciální remise). Výsledky a závěr: Metoda ProteoMiner umožnila na 2-DE mapách signifikantně zvýšit počet proteinových spotů ve srovnání s imunodepleční metodou MARS (Multiple Affinity Removal System). Mezi skupinami pacientů chemorezistentních a pacientů citlivých na léčbu bortezomibem bylo pomocí analýzy obrazu 2-DE gelů zjištěno 15 rozdílových proteinových spotů, které byly analyzovány hmotnostní spektrometrií. V těchto spotech bylo identifikováno 10 proteinů. Sedm proteinů vykazovalo signifikantně nižší hladinu proteinu ve skupině chemosenzitivních pacientů (sérový amyloid P, fibrinogen – gama řetězec, retinol-vazebný protein 4, komplement faktor C4-A, apolipoprotein E, karboxypeptidáza N, komplement faktoru H – příbuzný protein 1) a 3 proteiny vykazovaly signifikantně vyšší hladinu proteinu nebo byly detekovány pouze ve skupině chemosenzitivních pacientů (sérová paraoxonáza 1, alfa-1-antitrypsin a komplement faktor B).
Backgrounds: Recently, the term biomarker has become, especially in connection with the term clinical proteomics, one of the most frequent terms in the field of biomedical research. The aim of this work was to select an appropriate pre-fractionation method of blood plasma prior to a subsequent proteomic analysis of low-abundant fraction of proteins by two dimensional gel electrophoresis (2-DE) and mass spectrometry to improve the resolution of 2-DE maps and protein identification. Materials and Methods: First, we compared two prefractionation methods (MARS versus ProteoMiner) preceding 2-DE analysis using 10 blood plasma samples. Based on the results of the comparative experiments, low-abundant plasma protein fractions from 18 multiple myeloma patients treated with bortezomib were analyzed. Patients were divided into two groups: a group resistant to chemotherapy (9 patients – disease progression, stable disease) and a group with positive clinical response (9 patients – complete and partial remission). Results and Conclusion: Samples prefractioned by ProteoMiner method yielded 2-DE maps with a significantly increased number of detected protein spots, as compared to immunodepletion method MARS (Multiple Affinity Removal System). Between groups of chemoresistant and sensitive patients treated with bortezomib, 15 differently intense spots were revealed by image analysis. These spots were found to correspond to 10 proteins, as confirmed by mass spectrometry. Seven proteins had significantly lower protein level in the group of chemosensitive patients (serum amyloid P, fibrinogen – gamma chain, retinol-binding protein 4, complement factor C4-A, apolipoprotein E, carboxypeptidase N and complement factor H-related protein 1) and 3 proteins showed significantly higher levels of protein (or were only detected) in the group of chemosensitive patients (serum paraoxonase 1, alpha-1-antitrypsin and complement factor B).
- Klíčová slova
- elektroforéza gelová dvourozměrná, protein,
- MeSH
- 2D gelová elektroforéza metody využití MeSH
- biomedicínský výzkum MeSH
- bortezomib MeSH
- farmakoterapie MeSH
- financování organizované MeSH
- krevní plazma účinky léků MeSH
- krevní proteiny izolace a purifikace účinky léků MeSH
- kyseliny boronové aplikace a dávkování škodlivé účinky terapeutické užití MeSH
- lidé MeSH
- mnohočetný myelom farmakoterapie krev MeSH
- nádorové biomarkery krev MeSH
- proteomika metody MeSH
- pyraziny aplikace a dávkování škodlivé účinky terapeutické užití MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice metody využití MeSH
- statistika jako téma MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- srovnávací studie MeSH
The influence of the matrix solution, sample form and deposition technique on the quality MALDI-TOF mass spectra was examined and assessed with the aim to improve MALDI-TOF MS performance for the identification of microorganisms and to enable automatic spectra acquisition. It was observed that the use of matrix compounds ferulic and sinapinic acid may result in improved mass spectral features, in terms of signal resolution and S/N ratio, as compared to alpha-cyano-4-hydroxycinnamic acid, which was, on the other hand, found to be the only matrix compound that enabled fully automatic mass spectra acquisition. The robustness of the whole sample preparation procedure was then assessed on a set of 25 strains of four Acinetobacter species. Results showed reproducible detection of subtle mass spectral differences between strains belonging to the same species, although they do not confirm the possibility of reliable strain typing.
Acinetobacter genospecies (genomic species) 10 and 11 were described by Bouvet and Grimont in 1986 on the basis of DNA-DNA reassociation studies and comprehensive phenotypic analysis. In the present study, the names Acinetobacter bereziniae sp. nov. and Acinetobacter guillouiae sp. nov., respectively, are proposed for these genomic species based on the congruence of results of polyphasic analysis of 33 strains (16 and 17 strains of genomic species 10 and 11, respectively). All strains were investigated by selective restriction fragment amplification (i.e. AFLP) analysis rpoB sequence analysis, amplified rDNA restriction analysis and tDNA intergenic length polymorphism analysis, and their nutritional and physiological properties were determined. Subsets of the strains were studied by 16S rRNA gene sequence analysis and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS or had been classified previously by DNA-DNA reassociation. Results indicate that A. bereziniae and A. guillouiae represent two phenetically and phylogenetically distinct groups within the genus Acinetobacter. Based on the comparative analysis of housekeeping genes (16S rRNA and rpoB genes), these species together represent a monophyletic branch within the genus. Despite their overall phenotypic similarity, the ability to oxidize d-glucose and to grow at 38 degrees C can be used in the presumptive differentiation of these two species from each other: with the exception of three strains that were positive for only one test, A. bereziniae strains were positive for both tests, whereas A. guillouiae strains were negative in these tests. The strains of A. bereziniae originated mainly from human clinical specimens, whereas A. guillouiae strains were isolated from different environmental sources in addition to human specimens. The type strain of A. bereziniae sp. nov. is LMG 1003(T) (=CIP 70.12(T) =ATCC 17924(T)) and that of A. guillouiae sp. nov. is LMG 988(T) (=CIP 63.46( T) =ATCC 11171(T) =CCUG 2491(T)).
- MeSH
- Acinetobacter klasifikace genetika izolace a purifikace fyziologie MeSH
- analýza polymorfismu délky amplifikovaných restrikčních fragmentů MeSH
- druhová specificita MeSH
- fenotyp MeSH
- fylogeneze MeSH
- genotyp MeSH
- geny rRNA MeSH
- hybridizace nukleových kyselin MeSH
- infekce bakteriemi rodu Acinetobacter mikrobiologie MeSH
- infekce v ráně mikrobiologie MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- odpadní vody mikrobiologie MeSH
- ribozomální DNA analýza genetika MeSH
- RNA ribozomální 16S genetika MeSH
- sekvenční analýza DNA MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- techniky typizace bakterií MeSH
- zastoupení bazí MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The switch from aerobic to anaerobic respiration in the bacterium Paracoccus denitrificans is orchestrated by the action of three FNR-type transcription regulators FnrP, NNR and NarR, which are sensors for oxygen, nitric oxide and nitrite, respectively. In this work, we analyzed the protein composition of four strains (wild type, FnrP-, NNR- and NarR-mutant strains) grown aerobically, semiaerobically and semiaerobically in the presence of nitrate to discover the global role of FNR-family transcription regulators using proteomics, with data validation at the transcript and genome levels. Expression profiles were acquired using two-dimensional gel electrophoresis for 737 protein spots, in which 640 proteins were identified using mass spectrometry. The annotated 2-D proteome map provided the most comprehensive coverage of P. denitrificans proteome available to-date and can be accessed on-line at http://www.mpiib-berlin.mpg.de/2D-PAGE/. Our results revealed several types of regulation under the conditions tested: (1) FnrP-controlled regulation of nitrous oxide reductase, UspA and OmpW as confirmed at protein, transcript and DNA level (position of FNR boxes). (2) Proteins regulated via additional regulators, including proteins involved in NNR and NarR regulons: nitrate reductase beta-subunit, TonB-dependent receptors, nitrite reductase, a TenA-type transcription regulator, and an unknown protein with an alpha/beta hydrolase fold. (3) Proteins whose expression was affected mainly by the growth condition. This group contains SSU ribosomal protein S305 / sigma(54) modulation protein, and two short-chain reductase-dehydrogenase proteins.
- MeSH
- aerobióza fyziologie MeSH
- anaerobióza fyziologie MeSH
- bakteriální proteiny metabolismus MeSH
- dusitany metabolismus MeSH
- genetická transkripce fyziologie MeSH
- kyslík metabolismus MeSH
- oxid dusnatý metabolismus MeSH
- Paracoccus denitrificans fyziologie MeSH
- proteomika MeSH
- regulace genové exprese u bakterií fyziologie MeSH
- transkripční faktory metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Chemical communication in mammals involves globular lipocalins that protect and transport pheromones during their passage out of the body. Efficient communication via this protein - pheromone complex is essential for triggering multiple responses including aggression, mate choice, copulatory behaviour, and onset and synchronization of oestrus. The roles of lipocalins in communication were studied in many organisms and especially in mice (i.e. Mus musculus domesticus) which excrete Major Urinary Proteins (Mup) in excessive amounts in saliva and urine. Other mammals, however, often lack the genes for Mups or their expression is very low. Therefore, we aimed at characterization of candidate lipocalins in Myodes glareolus which are potentially linked to chemical communication. One of them is Aphrodisin which is a unique lipocalin that was previously described from hamster vaginal discharge and is known to carry pheromones stimulating copulatory behaviour in males. RESULTS: Here we show that Aphrodisin-like proteins exist in other species, belong to a group of Odorant Binding Proteins (Obp), and contrary to the expression of Aphrodisin only in hamster genital tract and parotid glands of females, we have detected these transcripts in both sexes of M. glareolus with the expression confirmed in various tissues including prostate, prepucial and salivary glands, liver and uterus. On the level of mRNA, we have detected three different gene variants. To assess their relevance for chemical communication we investigated the occurrence of particular proteins in saliva, urine and vaginal discharge. On the protein level we confirmed the presence of Obp2 and Obp3 in both saliva and urine. Appropriate bands in the range of 17-20 kDa from vaginal discharge were, however, beyond the MS detection limits. CONCLUSION: Our results demonstrate that three novel Obps (Obp1, Obp2, and Obp3) are predominant lipocalins in Myodes urine and saliva. On the protein level we have detected further variants and thus we assume that similarly as Major Urinary Proteins in mice, these proteins may be important in chemical communication in this Cricetid rodent.
- MeSH
- 2D gelová elektroforéza MeSH
- Arvicolinae genetika MeSH
- feromony genetika MeSH
- lipokaliny genetika sekrece MeSH
- moč chemie MeSH
- molekulární sekvence - údaje MeSH
- proteiny genetika MeSH
- receptory pachové genetika sekrece MeSH
- sekvence aminokyselin MeSH
- sekvenční seřazení MeSH
- sliny chemie MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- stanovení celkové genové exprese MeSH
- tandemová hmotnostní spektrometrie MeSH
- vagina chemie sekrece MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Výrazný rozvoj analytické instrumentace a metodických přístupů během posledních dvou desetiletí významně rozšířil možnosti studia proteinů v živých systémech. Analýza tzv. proteomu poskytuje neustále se prohlubující náhled do biologických procesů na úrovni kvalitativních a kvantitativních změn proteinového složení v souvislosti s fyziologickými a patologickými stavy organizmu, čímž nabízí možnost jejich lepšího pochopení a stává se přínosem pro vývoj a validaci diagnostických a terapeutických přístupů. Proto se na studium lidského proteomu v současné době soustřeďuje i pozornost odborné veřejnosti v oblasti onkologie. V tomto článku jsou vysvětleny principy nejvyužívanějších proteomických metod (gelová elektroforéza, kapalinová chromatografie, hmotnostně spektrometrická analýza, čipové technologie) a uvedeny příklady jejich využití v oblasti onkologických, resp. hematoonkologických onemocnění.
The rapid development of analytical instrumentation and methodical approaches in the course of the last two decades has significantly extended the possibilities of studying proteins in living systems. Proteomic analysis provides ever deeper insights into the molecular nature of biological processes in terms of qualitative and quantitative changes in protein composition in connection with the physiological and pathological states of the organism. Thus, proteomic analysis contributes to a better understanding of these processes and becomes a tool for the development and validation of diagnostic and therapeutic approaches. Thanks to recent achievements, the attention of cancer specialists is more and more focused on human proteome research. In this brief review we explain the principles of widely used proteomic techniques (gel electrophoresis, liquid chromatography, mass spectrometry analysis, protein array technologies) and show examples of their application in oncology, namely hematooncological diseases.
