Application of electron paramagnetic resonance spectroscopy in the study of albumin conformational changes by spin-labeling method

Серумски албумин је најзаступљенији протеин у крвној плазми, који има више важних физиолошких функција, због чега је детаљно проучаван разним техникама. Једна од техника коришћених за проучавање конформационих промена албумина и његовог капацитета за везивање разних супстанција је електронска парамагнетна резонантна (ЕПР) спектроскопија у спрези са спинским обележавањем. У ту сврху је коришћено неколико различитих спинских обележивача, међу којима су спински обележене масне киселине, с обзиром на то да је албумин главни транспортер масних киселина. Други често коришћени обележивач је 3-малеимидо проксил (5-МСЛ) који се ковалентно везује за слободан цистеин. У овој докторској дисертацији две спински обележене стеаринске киселине које садрже парамагнетну доксил групу (група од које потиче ЕПР сигнал) на петом (5-доксил стеаринска киселина, 5-ДС) и на шеснаестом (16-доксил стеаринска киселина, 16-ДС) угљениковом атому метиленског ланца су коришћене за проучавање конформационих промена у хуманом серумском албумину (ХСА). Осим тога, конфомационе промене у говеђем серумском албумину (БСА) су проучаване обележавањем БСА са 5-МСЛ. Конформационе промене су биле индуковане варирањем температуре и pH, додавањем етанола, везивањем неколико лиганада типичних за албумин (масне киселине и лекови) и излагањем дејству јаких оксидујућих агенаса, водоник-пероксиду и супероксидном анјонском радикалу. Додатни циљ ове дисертације је био да се испита да ли 5-МСЛ може да се користи за детекцију суптилних конформационих промена узрокованих везивањем лиганада, с обзиром на то да се 5-МСЛ налази у ригидној средини када се веже за једини слободан цистеин у БСA који се налази на позицији 34 у аминокиселинској секвенцији (Cys-34). Резултати ове дисертације показују да су доксил групе обележивача 5-ДС и 16-ДС (које дају информације о променамa у својој околини), лоциране на различитим местима у ХСА. Наиме, доксил група 5-ДС се налази у хидрофобној унутрашњости ХСА, док је за 16-ДС лоцирана близу површине ХСА или чак пролази кроз њу. Стога, ова два једињења дају информације са различитих места на молекулу ХСА. Такође је уочено да ЕПР спектри оба ДС обележивача везаних за ХСА могу да се разложе на компоненте које одговарају јако (ЈВ), слабо везаном (СВ) и невезаном (НВ) обележивачу. За 5-МСЛ везан за БСА, показано је да се ЕПР спектри састоје из две компоненте, ЈВ и СВ, које потичу од 5-МСЛ везаног за Cys-34 и амино групе, респективно...Serum albumin is the most abundant blood plasma protein, which has been thoroughly studied using a plethora of techniques, due to its multiple important physiological roles. One of the methods used to monitor albumin conformational changes and binding capacity is electron paramagnetic resonance (EPR) spin-labeling methodology. For this purpose, albumin has been studied using various spin-labels, among which spin-labeled fatty acids have been the most commonly employed, since albumin is the main carrier of fatty acids. The other frequently used spin-label is 3- maleimido-proxyl (5-MSL) which binds covalently to the free cysteine residues. In this doctoral dissertation, two spin-labeled stearic acids containing a paramagnetic doxyl group (the EPR-active group) attached at fifth (5-doxyl-stearic acid, 5-DS) and sixteenth (16-doxyl stearic acid, 16-DS) carbon atom of the methylene chain, were used to study the conformational changes of human serum albumin (HSA). Furthermore, conformational changes of bovine serum albumin (BSA) were studied by labeling BSA with 5-MSL. The conformational changes were induced by varying temperature and pH, addition of ethanol, binding several typical ligands (fatty acids and drugs) and exposure to strong oxidizing agents, hydrogen peroxide and superoxide anion radical. The additional goal of this dissertation was to reveal whether 5-MSL can be used to track subtle conformational changes arising from ligand binding, since 5- MSL is located in the fairly rigid environment when bound to the single free cysteine residue in BSA, located at the position 34 in amino acid sequence (Cys-34). The results from this study show that the doxyl groups (group which reports the changes in its environment) of 5-DS and 16-DS, are located at different sites in HSA. Namely, doxyl group of 5-DS is located in the hydrophobic interior, while for 16-DS it is close to, or even protrudes the surface of HSA. Hence, these two compounds give information from the different locations in HSA molecule. It was also observed that EPR spectra of both DS spin-labels bound to HSA could be decomposed into components corresponding to the strongly (SB), weakly bound (WB) and unbound (UB) label. In case of 5-MSL bound to BSA, it was shown that the corresponding EPR spectra consist of SB and WB component which originate from 5-MSL bound to Cys-34 and amino groups, respectively..

Lacrimal hyposecretion with women at videoterminals

U toku sveobuhvatnog pregleda radnica zaposlenih na videoterminalima uz obavezan okulistički pregled učinili smo i test ispitivanja sekrecije suza. Ispitivanja su obavljena na tri skupine po 100 radnica, i to prvoj gdje su bile radnice s punim radnim vremenom (8 sati) na terminalu, drugoj radnice s pola ili manje od pola radnog vremena (vrijeme provedeno za terminalom nije bilo manje od 2 sata na dan neprekidno) i trećoj, kontrolnoj, radnice koje borave u prostorijama gdje su videoterminali, ali rade neki drugi službenički posao. Hiposekrecija je najčešće bila prisutna kod radnica s punim radnim vremenom, i to kod 20% ispitanica. Statistički značajna razlika postoji između prve i druge dvije skupine ispitanica, dok između druge i kontrolne skupine nema značajne razlike.Tests of vision and lacrimal secretion were performed as part of a general medical check-up among women employed at videoterminals in a bank. The tests were taken by three groups of 100 employees each. The first group consisted of full-time (eight hours) employees, the second of those working part-time (the time spent at a videoterminal daily was never less than two hours of continuous work) and a third group of control subjects having different clerical jobs but working in the premises with videoterminals. Diminished lacrimal secretion was most often present among full-time workers (20 per cent). A statistically significant difference in lacrimal secretion was observed between the first and the second group of examinees. There was no significant difference between the second group and the controls

Assessment of possible association between rs378854 and prostate cancer risk in the Serbian population

Prostate cancer (PCa) is the second most commonly diagnosed cancer among men worldwide. Despite its high incidence rate, the molecular basis of PCa onset and its progression remains little understood. Genome-wide association studies (GWAS) have greatly contributed to the identification of single nucleotide polymorphisms (SNP) associated with PCa risk. Several GWAS identified 8q24 as one of the most significant PCa-associated regions. The aim of this study was to evaluate the association of SNP rs378854 at 8q24 with PCa risk in the Serbian population. The study population included 261 individuals diagnosed with PCa, 257 individuals diagnosed with benign prostatic hyperplasia (BPH) and 106 healthy controls. Data quality analysis yielded results showing deviations from Hardy-Weinberg equilibrium in groups of PCa patients and BPH patients as well as in the control group. There was no significant association between alleles and genotypes of the genetic variant rs378854 and PCa risk in the Serbian population. [Projekat Ministarstva nauke Republike Srbije, br. 173016

Clinical pattern of ocular toxoplasmosis treated in a referral centre in Serbia

Purpose To analyze the clinical pattern of ocular toxoplasmosis (OT) in a referral centre in Serbia. Patients and methods The medical records of consecutive patients admitted for OT to the single referral centre for uveitis in Serbia between 2006 and 2010 were retrospectively analyzed. OT was diagnosed on the basis of typical fundus lesions and positive serology for Toxoplasma. Results In a total of 457 uveitis patients, OT was the third leading cause, with 59 patients (12.9%). Most OT cases (73%) were monocular. An active primary retinal lesion was observed in 36% and recurrent OT in 64% patients. Localization of lesions was central/paracentral (44%), juxtapapillar (27%), peripheral (19%), and multifocal (10%). Other ocular manifestations of inflammation included vitritis (44%), anterior uveitis (19%), and retinal vasculitis (10%). Complications included choroidal neovascularization in two and exudative retinal detachment with cataract, glaucoma, and cystoid macular oedema in one patient each. The detection of Toxoplasma-specific IgM antibodies in a single patient indicates a low rate of OT concomitant with acute infection. After treatment, the mean best-corrected visual acuity (BCVA) increased significantly. However, 14 (24%) patients ended up legally blind in the affected eye, of which 2 (3%) with bilateral blindness, all with a very poor BCVA (0.047 +/- 0.055) at presentation. Visual impairment and treatment outcome were both associated with central localization of lesions (P lt 0.0001 and P = 0.006, respectively). Conclusion OT is a significant cause of posterior uveitis in Serbia. Patients should be aware of the recurring nature of OT and react immediately if symptoms occur. Eye (2012) 26, 723-728; doi: 10.1038/eye.2012.20; published online 24 February 201

Status and New Data of the Geochemical Determination of the pp-Neutrino Flux by LOREX

LOREX LORandite EXperiment addresses the determination of the solar pp neutrino flux during the last four million years by exploiting the reaction 205 Tl ν e → 205 Pb e − with an incomparably low-energy threshold of 50 keV for the capture of solar neutrinos. The ratio of 205 Pb/ 205 Tl atoms in the Tl-bearing mineral lorandite provides, if corrected for the cosmic-ray induced background, the product of the flux of solar neutrinos and their capture probability by 205 Tl, averaged over the age of lorandite. To get the mean solar neutrino flux itself, four problems have to be addressed: 1 the geological age of lorandite, 2 the amount of background cosmicray-induced 205 Pb atoms which strongly depends on the erosion rate of the lorandite-bearing rocks, 3 the capture probability of solar neutrinos by 205 Tl and 4 the extraction of lorandite and the appropriate technique to "count" the small number of 205 Pb atoms in relation to the number of 205 Tl atoms. This paper summarizes the status of items 1 age and 3 neutrino capture probability and presents in detail the progress achieved most recently concerning the items 2 background/erosion and 4 "counting" of 205 Pb atoms in lorandite

Corrigendum to "European contribution to the study of ROS:A summary of the findings and prospects for the future from the COST action BM1203 (EU-ROS)" [Redox Biol. 13 (2017) 94-162]

The European Cooperation in Science and Technology (COST) provides an ideal framework to establish multi-disciplinary research networks. COST Action BM1203 (EU-ROS) represents a consortium of researchers from different disciplines who are dedicated to providing new insights and tools for better understanding redox biology and medicine and, in the long run, to finding new therapeutic strategies to target dysregulated redox processes in various diseases. This report highlights the major achievements of EU-ROS as well as research updates and new perspectives arising from its members. The EU-ROS consortium comprised more than 140 active members who worked together for four years on the topics briefly described below. The formation of reactive oxygen and nitrogen species (RONS) is an established hallmark of our aerobic environment and metabolism but RONS also act as messengers via redox regulation of essential cellular processes. The fact that many diseases have been found to be associated with oxidative stress established the theory of oxidative stress as a trigger of diseases that can be corrected by antioxidant therapy. However, while experimental studies support this thesis, clinical studies still generate controversial results, due to complex pathophysiology of oxidative stress in humans. For future improvement of antioxidant therapy and better understanding of redox-associated disease progression detailed knowledge on the sources and targets of RONS formation and discrimination of their detrimental or beneficial roles is required. In order to advance this important area of biology and medicine, highly synergistic approaches combining a variety of diverse and contrasting disciplines are needed

New test of modulated electron capture decay of hydrogen-like 142Pm ions: Precision measurement of purely exponential decay

An experiment addressing electron capture (EC) decay of hydrogen-like 142Pm60+ions has been conducted at the experimental storage ring (ESR) at GSI. The decay appears to be purely exponential and no modulations were observed. Decay times for about 9000 individual EC decays have been measured by applying the single-ion decay spectroscopy method. Both visually and automatically analysed data can be described by a single exponential decay with decay constants of 0.0126(7)s−1for automatic analysis and 0.0141(7)s−1for manual analysis. If a modulation superimposed on the exponential decay curve is assumed, the best fit gives a modulation amplitude of merely 0.019(15), which is compatible with zero and by 4.9 standard deviations smaller than in the original observation which had an amplitude of 0.23(4)

European contribution to the study of ROS: A summary of the findings and prospects for the future from the COST action BM1203 (EU-ROS).

The European Cooperation in Science and Technology (COST) provides an ideal framework to establish multi-disciplinary research networks. COST Action BM1203 (EU-ROS) represents a consortium of researchers from different disciplines who are dedicated to providing new insights and tools for better understanding redox biology and medicine and, in the long run, to finding new therapeutic strategies to target dysregulated redox processes in various diseases. This report highlights the major achievements of EU-ROS as well as research updates and new perspectives arising from its members. The EU-ROS consortium comprised more than 140 active members who worked together for four years on the topics briefly described below. The formation of reactive oxygen and nitrogen species (RONS) is an established hallmark of our aerobic environment and metabolism but RONS also act as messengers via redox regulation of essential cellular processes. The fact that many diseases have been found to be associated with oxidative stress established the theory of oxidative stress as a trigger of diseases that can be corrected by antioxidant therapy. However, while experimental studies support this thesis, clinical studies still generate controversial results, due to complex pathophysiology of oxidative stress in humans. For future improvement of antioxidant therapy and better understanding of redox-associated disease progression detailed knowledge on the sources and targets of RONS formation and discrimination of their detrimental or beneficial roles is required. In order to advance this important area of biology and medicine, highly synergistic approaches combining a variety of diverse and contrasting disciplines are needed.The EU-ROS consortium (COST Action BM1203) was supported by the European Cooperation in Science and Technology (COST). The present overview represents the final Action dissemination summarizing the major achievements of COST Action BM1203 (EU-ROS) as well as research news and personal views of its members. Some authors were also supported by COST Actions BM1005 (ENOG) and BM1307 (PROTEOSTASIS), as well as funding from the European Commission FP7 and H2020 programmes, and several national funding agencies

Cervikobrahijalni sindrom - rad i rdna nesposobnost

The subjects in the study were 114 persons receiving a disability pension or referred for disability assessment. They were placed in two groups of 57 persons each, one with marked cervicobrachial syndrome and the other without. The latter group was chosen by the method of equivalent pairs with regard to sex and age. All subjects underwent a clinical examination and a standardized questionnaire was completed. The questionnaire pertained to the state of health, focussing on the amount and type of physical burdening at the workplace and on activities outside the workplace. Sixty-five percent of the subjects were aged from 51 to 60 years. A statistically significant difference was established between earlier occupation and cervicobrachial syndrome. Cervicobrachial syndrome was found in 31.6% of unskilled workers and 12.3% of the controls (P<0.05). A forced body posture during work was recorded in 74% or the subjects with the syndrome and in 50% of the control subjects (P<0.05). Repetitive movements at work were reported by a large number of subjects with cervicobrachial syndrome (71.9:49.1%; P<0.05) who also claimed to strain the arms (84.2:61.4%; P<0.05) and burden the cervical spine (68.5:40.4%; P<0.05) during household activities to a significantly greater extent than the controls. A possible preventive approach to the occurrence and progression of cervicobrachial syndrome is discussed.Istraživanjem je obuhvaćeno 114 ispitanika koji su odabrani iz skupine invalidskih umirovljenika te bolesnika koji su bili na zdravstvenoj obradi za utvrđivanje invalidnosti. Ispitivanu je skupinu činilo 57 invalidskih umirovljenika s jasno izraženim cervikobrahijalnim sindromom. Poredbena skupina, formirana metodom ekvivalentnih parova s obzirom na spol i dob, bili su bolesnici kojima se ovaj sindrom sa sigurnošću mogao isključiti. Za sve je ispitanike uz klinički pregled ispunjen i standardizirani upitnik u kojem su uz podatke o zdravstvenom stanju bila pitanja o opterećenjima na radnom mjestu, kao i aktivnostima izvan rada. Većina ispitanika (65%) bila je u dobi između 51 i 60 godina, Utvrđena je statistički značajna razlika u zastupanosti cervikobrahijalnog sindroma s obzirom na ranije zanimanje. U ispitivanoj skupini nekvalificirani radnici bili su zastupani s 31,6%, a u poredbenoj s 12,3% (P<0,05). Prisilan položaj tijela pri radu zabilježen je u 74% ispitanika s cervikobrahijalnim sindromom te u 50% ispitanika bez ijednog simptoma ovog sindroma (P<0,05). Jednake pokrete pri radu također je navodio veći broj ispitanika s cervikobrahijalnim sindromom