Determination Of Protein Content Research Articles

Reduction of lung inflammation, oxidative stress and apoptosis by the PDE4 inhibitor roflumilast in experimental model of acute lung injury.

Damage of alveolar-capillary barrier, inflammation, oxidative injury, and lung cell apoptosis represent the key features of acute lung injury (ALI). This study evaluated if selective phosphodiesterase (PDE)-4 inhibitor roflumilast can reduce the mentioned changes in lavage-induced model of ALI. Rabbits with ALI were divided into 2 groups: ALI without therapy (A group) and ALI treated with roflumilast i.v. (1 mg/kg; A+R group). One group of healthy animals without ALI served as ventilated controls (C group). All animals were oxygen-ventilated for further 4 h. At the end of experiment, total and differential counts of cells in bronchoalveolar lavage fluid (BALF) and total and differential counts of white blood cells were estimated. Lung edema formation was assessed from determination of protein content in BALF. Pro-inflammatory cytokines (TNFalpha, IL-6 and IL-8) and markers of oxidation (3-nitrotyrosine, thiobarbituric-acid reactive substances) were detected in the lung tissue and plasma. Apoptosis of lung cells was investigated immunohistochemically. Treatment with roflumilast reduced leak of cells, particularly of neutrophils, into the lung, decreased concentrations of cytokines and oxidative products in the lung and plasma, and reduced lung cell apoptosis and edema formation. Concluding, PDE4 inhibitor roflumilast showed potent anti-inflammatory actions in this model of ALI.

Sparse NIR optimization method (SNIRO) to quantify analyte composition with visible (VIS)/near infrared (NIR) spectroscopy (350 nm-2500 nm)

Visual-Near-Infra-Red (VIS/NIR) spectroscopy has led the revolution in high-throughput phenotyping methods used to determine chemical and structural elements of organic materials. In the current state of the art, spectrophotometers used for imaging techniques are either very expensive or too large to be used as a field-operable device. In this study we developed a Sparse NIR Optimization method (SNIRO) that selects a pre-determined number of wavelengths that enable quantification of analytes in a given sample using linear regression. We compared the computed complexity time and the accuracy of SNIRO to Marten's test, to forward selection test and to LASSO all applied to the determination of protein content in corn flour and meat and octane number in diesel using publicly available datasets. In addition, for the first time, we determined the glucose content in the green seaweed Ulva sp., an important feedstock for marine biorefinery. The SNIRO approach can be used as a first step in designing a spectrophotometer that can scan a small number of specific spectral regions, thus decreasing, potentially, production costs and scanner size and enabling the development of field-operable devices for content analysis of complex organic materials.

Effect of Protein Supplements on Renal Function and Lipid Profile

Background: Protein supplements are one of the most frequently dietary supplements used by athletes and non-athletes in Saudi Arabia.
 Objective: To assess the knowledge, attitude and practice of Saudi population toward the protein supplements, examine the relationships of practice with education level, effect of protein on kidneys and lipid profile, assess the types and source of protein supplements, assess the correct use of protein supplements according to exercise intensity and determine protein content and heavy metals in protein supplements.
 Methods: A national cross-sectional survey was conducted during October-December 2017 on protein supplements users in Saudi Arabia. Moreover, samples of the best selling protein supplements were analyzed to determined total protein and heavy metals content.
 Results: A total of 398 filled protein supplements user questionnaire. The wrong practice of use associated with the effect of protein supplements on renal function and lipid profile.The percentage of protein content in isolated whey protein (68.18%) whiles by Kjeldhl analysis (80.55%). Concentrated whey protein contain (72%) and by kjeldhl analysis (77.2%). Hydrolyzed collagen containing (100%) protein and by kjeldhl analysis (91.25%).The amount of heavy metals in isolated whey protein contains arsenic (83 μg/kg), mercury (<10 μg/kg) and lead (18 μg/kg), while hydrolyzed collagen sample contains arsenic (101 μg/kg), mercury (<10 μg/kg) and lead (49 μg/kg).
 Conclusion: The current study reveals an evidence for lacking of the awareness, negative attitude and abuse of protein supplements, demonstrates necessity to further education regarding the use and purchasing of protein supplements to ensure the quality of these products.

Undergraduate laboratory experiment on determination of total protein content in milk powder by moving reaction boundary titration.

Laboratory exercises focused on protein quantification are frequently conducted in traditional undergraduate biochemistry laboratory curriculum. The laboratory course described here is designed to provide students with experience in measurement of protein content in milk powder by moving reaction boundary titration (MRBT), a new rapid technique for total protein content determination in milk. In addition, this approach is weakly influenced by nonprotein nitrogen reagents such as melamine and urea. The course was done as three weekly laboratory exercises. First, students established a standard curve for milk protein concentration by MRBT method. Then, students investigated the influence of nonprotein nitrogen reagents on MRBT method. Finally, students made a comparison among three different protein quantification methods (MRBT, Biuret, and Kjeldahl method). From the experiments, students grasped the concept and advantages of MRBT and deepened the understanding of protein quantification. This course offer students the opportunity to be exposed to an advanced technique, which may have practical significance to their future study and work in the life science field. © 2018 International Union of Biochemistry and Molecular Biology, 46(6):644-651, 2018.

Determination of aflatoxin, protein and fat content of ‘Tombul’ hazelnut under different drying conditions used by farmers in Giresun province of Turkey

This research was carried out on 'Tombul' hazelnut in 2009 in Giresun province of Turkey. It was aimed to investigate the aflatoxin, protein and total fat contents of hazelnut under 3 different drying conditions, in three different orchards, used by farmers. In the study, jute, polyethylene and concrete conditions were used for drying. The results showed that there was no aflatoxin contamination for all groups. The protein contents of the samples differed according to the drying conditions and interaction, and changed from 15.71 (polyethylene) to 16.60% (concrete). There was no difference between jute and polyethylene. The highest protein value (16.87%) was observed for concrete × first orchard samples, and the lowest for polyethylene × second orchard as interaction. The fat contents of the samples differed according to the drying conditions and orchards, and changed from 58.37 (jute) to 62.17% (polyethylene) as drying conditions. There was no difference between jute and concrete. The fat contents of the samples in the different orchards were 58.70, 59.69, and 61.18% for 1, 2, and 3, respectively.

FTIR-ATR determination of protein content to evaluate whey protein concentrate adulteration

We evaluated the potential application of Fourier-Transformed Infrared spectroscopy using Attenuated Total Reflectance (FTIR-ATR) to characterize and detect adulteration of whey protein concentrate (WPC). Samples were adulterated by substituting WPC by milk whey powder (MWP). Principal Component Analysis (PCA) was used to characterize the spectra. Partial Least Squares (PLS) regressions were applied to model and predict the protein content and the amount in grams of WPC and MWP in the samples. In WPC, Amide I and II bands showed decreasing trend with the protein content of the samples. The carbohydrate region between 1160 and 1000 cm−1 showed spectra with opposite behavior of the Amide bands, evincing the ν CO peak (1030 cm−1) that increases as MWP was added. In PCA, PC1 (64.5%) and PC2 (18.8%) described above 80% of total variance of the spectra. PC1 variance was most influenced by Amide I, II and carbohydrate regions. In all cases, PLS showed low prediction errors, high precision and coefficients of determination of the global fit above 0.99. Therefore, the association of FTIR-ATR spectroscopy with multivariate approaches revealed strong potential to detect adulteration in nutritional supplements and high accuracy to predict simultaneously the protein content and mass of WPC and MWP added.

Oxygen conserving mitochondrial adaptations in the skeletal muscles of breath hold divers.

BackgroundThe performance of elite breath hold divers (BHD) includes static breath hold for more than 11 minutes, swimming as far as 300 m, or going below 250 m in depth, all on a single breath of air. Diving mammals are adapted to sustain oxidative metabolism in hypoxic conditions through several metabolic adaptations, including improved capacity for oxygen transport and mitochondrial oxidative phosphorylation in skeletal muscle. It was hypothesized that similar adaptations characterized human BHD. Hence, the purpose of this study was to examine the capacity for oxidative metabolism in skeletal muscle of BHD compared to matched controls.MethodsBiopsies were obtained from the lateral vastus of the femoral muscle from 8 Danish BHD and 8 non-diving controls (Judo athletes) matched for morphometry and whole body VO2max. High resolution respirometry was used to determine mitochondrial respiratory capacity and leak respiration with simultaneous measurement of mitochondrial H2O2 emission. Maximal citrate synthase (CS) and 3-hydroxyacyl CoA dehydrogenase (HAD) activity were measured in muscle tissue homogenates. Western Blotting was used to determine protein contents of respiratory complex I-V subunits and myoglobin in muscle tissue lysates.ResultsMuscle biopsies of BHD revealed lower mitochondrial leak respiration and electron transfer system (ETS) capacity and higher H2O2 emission during leak respiration than controls, with no differences in enzyme activities (CS and HAD) or protein content of mitochondrial complex subunits myoglobin, myosin heavy chain isoforms, markers of glucose metabolism and antioxidant enzymes.ConclusionWe demonstrated for the first time in humans, that the skeletal muscles of BHD are characterized by lower mitochondrial oxygen consumption both during low leak and high (ETS) respiration than matched controls. This supports previous observations of diving mammals demonstrating a lower aerobic mitochondrial capacity of the skeletal muscles as an oxygen conserving adaptation during prolonged dives.

An intrinsic fluorescence method for the determination of protein concentration in vaccines containing aluminum salt adjuvants

Determination of protein concentration in vaccines containing aluminum salt adjuvant typically necessitates desorption of the protein prior to analysis. Here we describe a method based on the intrinsic fluorescence of tyrosine and tryptophan that requires no desorption of proteins. Adjuvanted formulations of three model Bordetella pertussis antigens were excited at 280 nm and their emission spectra collected from 290 to 400 nm. Emission spectra of protein antigens in the presence of aluminum salt adjuvants were able to be detected, the effects of adjuvants on the spectra were analyzed, and linear regressions were calculated. The fluorescence method proved to be very sensitive with a limit of quantification between 0.4 and 4.4 µg/mL and limit of linearity between 100 and 200 µg/mL, across the formulations tested. The fluorescence method was found to be influenced by adjuvant presence, type of adjuvant, adjuvant concentration, buffer and pH conditions. The method also demonstrated ability to monitor the percent adsorption of antigens to the adjuvants. Furthermore, intrinsic fluorescence showed good correlation with micro-Kjeldahl elemental assay in quantifying protein concentration. Being a non-invasive, quick and sensitive method, intrinsic fluorescence has the potential to be utilized as a high throughput tool for vaccine development and conceivably implemented in-line, using in-line fluorimeters, to monitor antigen concentration during formulation processing.

Effect of thoracic paravertebral block combined with general anesthesia on postoperative cognitive dysfunction in elderly patients undergoing pulmonary lobectomy

Objective To evaluate the effect of thoracic paravertebral block combined with general anesthesia on postoperative cognitive dysfunction (POCD) in elderly patients undergoing pulmonary lobectomy. Methods A total of 120 elderly patients of both sexes, aged 65-81 yr, of American Society of Anesthesiologists physical status Ⅱ or Ⅲ, with New York Heart Association Ⅰor Ⅱ, were divided into 3 groups (n=40 each) using a random number table method: anesthesia group (group GA), epidural block combined with general anesthesia group (group EG), and paravertebral block combined with general anesthesia group (group PG). In group PG, paravertebral block was performed under ultrasound guidance, 0.25% ropivacaine 20 ml was injected after the paravertebral catheter was placed, and anesthesia was induced after confirming the plane of block.In group EG, epidural block was performed with 2% lidocaine 3 ml after epidural puncture was successfully performed at L6, 7 interspace, epidural 0.375% ropivacaine 8-15 ml was intermittently injected, and anesthesia was induced after confirming the height of block.Anesthesia was induced with IV midazolam 0.05-0.10 mg/kg, etomidate 0.3 mg/kg, sufentanil 0.4 μg/kg and rocuronium 0.6 mg/kg.The patients were tracheally intubated and mechanically ventilated.The development of POCD was recorded at 1 day before operation and 7 days after operation.Blood samples were collected from the internal jugular vein before anesthesia, at 15 min after skin incision and at 7 days after operation for determination of serum adiponectin (ADP) and S-100β protein concentrations. Results Compared with group GA, the incidence of POCD was significantly decreased, and the serum S-100β protein concentrations were decreased and serum ADP concentrations were increased at 15 min after skin incision and 7 days after operation in PG and EG groups (P<0.05). Compared with group EG, the incidence of POCD was significantly decreased, and the serum S-100β protein concentrations were increased and serum ADP concentrations were decreased at 15 min after skin incision and 7 days after operation in group PG (P<0.05). Conclusion Thoracic paravertebral block combined with general anesthesia induces better efficacy in decreasing the occurrence of POCD than general anesthesia alone or combination of epidural block and general anesthesia in elderly patients undergoing pulmonary lobectomy, which is related to the decreased concentrations of blood ADP in elderly patients undergoing pulmonary lobectomy. Key words: Nerve block; Pneumonectomy; Cognition disorders; Aged

Characterizing the Surface Coverage of Protein-Gold Nanoparticle Bioconjugates.

Functional enzyme-nanoparticle bioconjugates are increasingly important in biomedical and biotechnology applications such as drug delivery and biosensing. Optimization of the function of such bioconjugates requires careful control and characterization of their structures and activity, but current methods are inadequate for this purpose. A key shortcoming of existing approaches is the lack of an accurate method for quantitating protein content of bioconjugates for low (monolayer) surface coverages. In this study, an integrated characterization methodology for protein-gold nanoparticle (AuNP) bioconjugates is developed, with a focus on site-specific attachment and surface coverage of protein on AuNPs. Single-cysteine-containing mutants of dihydrofolate reductase are covalently attached to AuNPs with diameters of 5, 15, and 30 nm, providing a range of surface curvature. Site-specific attachment to different regions of the protein surface is investigated, including attachment to a flexible loop versus a rigid α helix. Characterization methods include SDS-PAGE, UV-vis spectrophotometry, dynamic light scattering, and a novel fluorescence-based method for accurate determination of low protein concentration on AuNPs. An accurate determination of both protein and AuNP concentration in conjugate samples allows for the calculation of the surface coverage. We find that surface coverage is related to the surface curvature of the AuNP, with a higher surface coverage observed for higher surface curvature. The combination of these characterization methods is important for understanding the functionality of protein-AuNP bioconjugates, particularly enzyme activity.

Light-Driven Chloride Transport Kinetics of Halorhodopsin

Despite growing interest in light-driven ion pumps for use in optogenetics, current estimates of their transport rates span two orders of magnitude due to challenges in measuring slow transport processes and determining protein concentration and/or orientation in membranes in vitro. In this study, we report, to our knowledge, the first direct quantitative measurement of light-driven Cl− transport rates of the anion pump halorohodopsin from Natronomonas pharaonis (NpHR). We used light-interfaced voltage clamp measurements on NpHR-expressing oocytes to obtain a transport rate of 219 (± 98) Cl−/protein/s for a photon flux of 630 photons/protein/s. The measurement is consistent with the literature-reported quantum efficiency of ∼30% for NpHR, i.e., 0.3 isomerizations per photon absorbed. To reconcile our measurements with an earlier-reported 20 ms rate-limiting step, or 35 turnovers/protein/s, we conducted, to our knowledge, novel consecutive single-turnover flash experiments that demonstrate that under continuous illumination, NpHR bypasses this step in the photocycle.

Efficiency of photodynamic therapy and Avastin in treatment of corneal neovascularization in rabbit's eye

The aim of the present work is to evaluate the effect of photodynamic therapy (PDT) by using argon laser with rose bengal (Herbs) as a photosensitizer when compared to the Avastin for treatment of corneal neovascularization (CNV) in rabbits. Thirty-Nine New Zealand male rabbits aged 2.5 months and weighing 2.5-3 kg are selected from the animal house of research institute of ophthalmology, Giza, Egypt. Three rabbits are used as control, and 36 rabbits are subjected to corneal neovascularization (CNV) induced by 3 silk sutures at mid-stromal depth approximately 1 mm from the limbus. After four days, sutures are removed and slit lamp examinations are performed to determine the area of CNV. Rabbits with CNV are divided into 3 groups: (1) The first group left without any treatment(n=12); (2) The second group (n=12 rabbits) were generally anesthetized and rose bengal(C20H4Cl4I4O5) with a dose of 50 mg/Kg was administered by slow intravenous injection in the marginal ear vein. After 15 minutes, rabbit’s eyes were exposed to argon laser(Quantel-Medical, Vitra, France) in a pulsed mode at 532 nm, spot size of 50 μm, pulse duration of 0.2 ms and a power of 150-200 mW/cm 2.; (3) The third group are treated with subconjunctival injection of 25 mg Avastin (n=12). The rabbits are followed up for 4 weeks and 3 rabbits are selected and sacrificed weekly from each group. The corneas are isolated for determination of protein content, SDS-poly acrylamide gel electrophoresis, total antioxidant capacity (TAC) and total oxidative capacity (TOC). Final results show that avastin is almost as effective as photodynamic therapy.

Electro-responsive graphene oxide hydrogels for skin bandages: The outcome of gelatin and trypsin immobilization

A free radical polymerization method was adopted for the fabrication of hybrid hydrogel films based on acrylamide and polyethylene glycol dimethacrylate as plasticizing and crosslinking agents, respectively, to be employed as smart skin bandages. Electro-sensitivity, biocompatibility and proteolytic properties were conferred to the final polymer networks by introducing graphene oxide (0.5% w/w), gelatin or trypsin (10% w/w) in the polymerization feed. The physical chemical and mechanical characterization of hybrid materials was performed by means of determination of protein content, Raman spectroscopy, thermogravimetric analysis and measurement of tensile strength. The evaluation of both water affinity and curcumin release profiles (analyzed by suitable mathematical modelling) upon application of an external electric stimulation in the 0–48 voltage range, confirmed the possibility to modulate the release kinetics. Proper proteolytic tests showed that the trypsin enzymatic activity was retained by 80% upon immobilization. Moreover, for all samples, we observed a viability higher than 94% in normal human fibroblast cells (MRC-5), while a reduction of methicillin-resistant Staphylococcus aureus CFU mL−1 (90%) was obtained with curcumin loaded samples.

Grafting of Oligo(ethylene glycol)-Functionalized Calix[4]arene-Tetradiazonium Salts for Antifouling Germanium and Gold Surfaces.

Biosensors that can determine protein concentration and structure are highly desired for biomedical applications. For the development of such biosensors, the use of Fourier transform infrared (FTIR) spectroscopy with the attenuated internal total reflection (ATR) configuration is particularly attractive, but it requires appropriate surface functionalization of the ATR optical element. Indeed, the surface has to specifically interact with a target protein in close contact with the optical element and must display antifouling properties to prevent nonspecific adsorption of other proteins. Here, we report robust monolayers of calix[4]arenes bearing oligo(ethylene glycol) (oEG) chains, which were grafted on germanium and gold surfaces via their tetradiazonium salts. The formation of monolayers of oEGylated calix[4]arenes was confirmed by AFM, IR, and contact angle measurements. The antifouling properties of these modified surfaces were studied by ATR-FTIR spectroscopy and fluorescence microscopy, and the nonspecific absorption of bovine serum albumin was found to be reduced by 85% compared to that of unmodified germanium. In other words, the organic coating by oEGylated calix[4]arenes provides remarkable antifouling properties, opening the way for the design of germanium- or gold-based biosensors.

COMPARATIVE STUDIES OF EGG ALBUMIN FROM DIFFERENT SOURCES

This work describes the use of different methods to study egg white proteins from hen, duck and quail. For this purpose, protein content determination, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Urea-polyacrylamide gel electrophoresis (Urea-PAGE), amino acids composition and Fourier Transform Infrared (FTIR) Spectroscopy were used to compare egg white proteins from these poultry species. Protein content, ranging from 71.31 to 72.36%. Quail egg white showed the lowest protein concentration (71.31%). The highest protein concentration was observed in duck egg white, which reached 72.36%. The analysis by SDS-PAGE, egg white proteins presented a wide range of relative molecular masses. The main egg white proteins are ovalbumin, ovotransferrin, ovomucoid and lysozyme. The contents of the hydrophobic amino acids residues (Pro, Gly, Ala, Val, Ile, Leu, Phe) are 34.54%, 34.95% and 34.94% for egg albumin from hen, duck and quail, respectively. The migration in Urea-PAGE into cathode direction indicated that duck egg white protein and quail egg white protein were much faster than hen egg white protein referring to bigger positive charges. Also, the secondary structure of protein was observed by FTIR.

Effects of U50488H on postoperative cognitive dysfunction induced by cardiopulmonary bypass in rats

Objective To investigate the effect of U50488H on postoperative cognitive dysfunction induced by cardiopulmonary bypass (CPB) in rats. Methods Forty-eight adult male Sprague-Dawley rats, weighing 400-450 g, were divided into 3 groups (n=16 each) using a random number table: sham operation group (group S), CPB group and CPB plus U50488H group (group U). CPB was performed for 60 min in group CPB.U50488H 1.5 mg/kg was injected into the left lateral cerebral ventricle at 30 min prior to CPB, and then CPB was performed for 60 min in group U. Eight rats in each group were selected at 1 day after CPB, and venous blood samples were collected for determination of serum S100β protein, interleukin-1beta (IL-1β) and tumor necrosis factor-alpha (TNF-α) concentrations (by enzyme-linked immunosorbent assay). Then the rats were sacrificed and right hippocampi were removed for examination of the pathological changes after haematoxylin and eosin staining.Eight rats were selected from each group at 7 days after CPB for assessment of cognitive function. Results Compared with group S, the concentrations of serum S100β protein, IL-1β and TNF-α were significantly increased in group CPB, and the escape latency was prolonged, the number of crossing original platform was reduced, and the swimming distance in target quadrant and time of staying at target quadrant were shortened in CPB and U groups (P<0.05). Compared with group CPB, the concentrations of serum S100β protein, IL-1β and TNF-α were significantly decreased, the escape latency was shortened, the number of crossing original platform was increased, the swimming distance in target quadrant and time of staying at target quadrant were prolonged (P<0.05), and the pathological changes of hippocampal tissues were significantly attenuated in group U. Conclusion U50488H can mitigate the postoperative cognitive dysfunction induced by CPB in rats. Key words: Receptors, opioid, kappa; Cardiopulmonary bypass; Cognition disorders

Precise determination of protein extinction coefficients under native and denaturing conditions using SV-AUC.

The accurate determination of protein concentration is an important though non-trivial task during the development of a biopharmaceutical. The fundamental prerequisite for this is the availability of an accurate extinction coefficient. Common approaches for the determination of an extinction coefficient for a given protein are either based on the theoretical prediction utilizing the amino acid sequence or the photometric determination combined with a measurement of absolute protein concentration. Here, we report on an improved SV-AUC based method utilizing an analytical ultracentrifuge equipped with absorbance and Rayleigh interference optics. Global fitting of datasets helped to overcome some of the obstacles encountered with the traditional method employing synthetic boundary cells. Careful calculation of dn/dc values taking glycosylation and solvent composition into account allowed the determination of the extinction coefficients of monoclonal antibodies and an Fc-fusion protein under native as well as under denaturing conditions. An intra-assay precision of 0.9% and an accuracy of 1.8% compared to the theoretical value was achieved for monoclonal antibodies. Due to the large number of data points of a single dataset, no meaningful difference between the ProteomeLab XL-I and the new Optima AUC platform could be observed. Thus, the AUC-based approach offers a precise, convenient and versatile alternative to conventional methods like total amino acid analysis (AAA).

Brain natriuretic peptide serum levels in periodontitis.

N-terminal-pro-brain natriuretic peptide (NT-proBNP) is expressed during inflammation and considered to be a biomarker of cardiovascular disease (CVD). In the last decade, evidence has suggested that periodontitis is associated with CVD. However, little is known of the potential role of this peptide in periodontal disease. The aims of this study were to examine whether the levels of NT-proBNP in serum are increased in periodontal disease and if there is a relationship to severity of periodontitis. A case-control study was carried out, in which serum samples were collected from 40 patients with periodontitis and from 40 nonperiodontitis individuals. Full periodontal examination was performed in all subjects. Demographic information, and clinical and history of certain diseases were also recorded. Determinations of serum NT-proBNP and high-sensitivity C-reactive protein concentrations were carried out in an independent laboratory. The levels of NT-proBNP in serum were significantly higher in patients with periodontitis compared with controls (87.9pg/mL vs 29.9pg/mL, P<.0001). In addition, as periodontal disease progressed, the levels of NT-proBNP increased. Multivariable regression analysis showed that the periodontal inflamed surface area, a measure of periodontal inflammation and disease activity, was the only periodontal parameter significantly associated with elevated concentrations of NT-proBNP in serum (R2 =.777, P<.0001). In periodontitis, increased serum NT-proBNP levels are observed in comparison with individuals without periodontitis. Moreover, the greater the degree of periodontal destruction, the higher the levels of NT-proBNP in serum.

Hybrid structures based on gold nanoparticles and semiconductor quantum dots for biosensor applications.

BackgroundThe luminescence amplification of semiconductor quantum dots (QD) in the presence of self-assembled gold nanoparticles (Au NPs) is one of way for creating biosensors with highly efficient transduction.AimsThe objective of this study was to fabricate the hybrid structures based on semiconductor CdSe/ZnS QDs and Au NP arrays and to use them as biosensors of protein.MethodsIn this paper, the hybrid structures based on CdSe/ZnS QDs and Au NP arrays were fabricated using spin coating processes. Au NP arrays deposited on a glass wafer were investigated by optical microscopy and absorption spectroscopy depending on numbers of spin coating layers and their baking temperature. Bovine serum albumin (BSA) was used as the target protein analyte in a phosphate buffer. A confocal laser scanning microscope was used to study the luminescent properties of Au NP/QD hybrid structures and to test BSA.ResultsThe dimensions of Au NP aggregates increased and the space between them decreased with increasing processing temperature. At the same time, a blue shift of the plasmon resonance peak in the absorption spectra of Au NP arrays was observed. The deposition of CdSe/ZnS QDs with a core diameter of 5 nm on the surface of the Au NP arrays caused an increase in absorption and a red shift of the plasmon peak in the spectra. The exciton–plasmon enhancement of the QDs’ photoluminescence intensity has been obtained at room temperature for hybrid structures with Au NPs array pretreated at temperatures of 100°C and 150°C. It has been found that an increase in the weight content of BSA increases the photoluminescence intensity of such hybrid structures.ConclusionThe ability of the qualitative and quantitative determination of protein content in solution using the Au NP/QD structures as an optical biosensor has been shown experimentally.

Effect of Acetylation on Liquid‐Liquid Phase Separation and Amyloid Fibril Formation of Tau

The function of proteins depends on their phase behavior. When the concentration of a protein exceeds a critical value, it may undergo liquid‐liquid phase separation, leading to demixing of proteins from solution. The resulted liquid droplets are dense and protein‐rich. The microtubule associated protein tau can enrich tubulin by forming liquid droplets, which promotes the formation of microtubule bundles. On the other hand, aberrant aggregation of Tau into neurofibrillary tangles correlates with the progressive destruction of nerve cells. Therefore, studying the aggregating mechanism of tau and drug design targeting tau fibrils are critical for the therapy of Alzheimer's disease. Recent studies have shown that lysine acetylation plays an important role in modulating tau aggregation and microtubule assembly. Considering the potential correlations between liquid‐liquid phase separation and amyloid formation, here we investigated the effect of lysine acetylation on the phase behavior and function of tau in vitro. We used site‐directed mutagenesis, light scattering, thioflavin T florescence assay, and microrheology to study the liquid‐liquid phase separation and aggregation of tau. Consistent with previous observations, acetylation on K274/K321 (mimicked by K274Q/K321Q) significantly impaired tau aggregation whereas acetylation on K280/K281/K369 (mimicked by K280Q/K281Q/K369Q) significantly promoted tau filament formation as compared with the WT protein. Although acetylation on K274/K321 and K280/K281/K369 differentially affected tau aggregation, both acetylation patterns promoted liquid‐liquid phase separation of tau. Moreover, microrheology characterization and protein concentration determination showed that the droplets formed by acetylated tau were denser than the WT protein, suggesting that more intermolecular interactions were form upon lysine acetylation. These results show that lysine residues within the MT‐binding repeats regulate the phase behavior of tau via complex interactions.Support or Funding InformationThis work was supported by National Natural Science Foundation of China (Grant number: 21603121) and Hubei University of Technology.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.