Effects of intracellular calcium accumulation on proteins encoded by the major genes underlying amyotrophic lateral sclerosis

The aetiology of Amyotrophic Lateral Sclerosis (ALS) is still poorly understood. The discovery of genetic forms of ALS pointed out the mechanisms underlying this pathology, but also showed how complex these mechanisms are. Excitotoxicity is strongly suspected to play a role in ALS pathogenesis. Excitotoxicity is defined as neuron damage due to excessive intake of calcium ions (Ca2+) by the cell. This study aims to find a relationship between the proteins coded by the most relevant genes associated with ALS and intracellular Ca2+ accumulation. In detail, the profile of eight proteins (TDP-43, C9orf72, p62/sequestosome-1, matrin-3, VCP, FUS, SOD1 and profilin-1), was analysed in three different cell types induced to raise their cytoplasmic amount of Ca2+. Intracellular Ca2+ accumulation causes a decrease in the levels of TDP-43, C9orf72, matrin3, VCP, FUS, SOD1 and profilin-1 and an increase in those of p62/sequestosome-1. These events are associated with the proteolytic action of two proteases, calpains and caspases, as well as with the activation of autophagy. Interestingly, Ca2+ appears to both favour and hinder autophagy. Understanding how and why calpain-mediated proteolysis and autophagy, which are physiological processes, become pathological may elucidate the mechanisms responsible for ALS and help discover new therapeutic targets.

THROMBOSE-FIBROUS COMPLICATIONS OF SYSTEMIC ENDOTHELIUM DAMAGE AT COVID-19

The analysis of literature data and our own research of lungs tissues of the persons who died owing to COVID-19 caused fibrosis testify to participation in this pathology of a cascade of disturbances of molecular and cellular levels. Viral damage to endothelial cells causes systemic damage to the vascular glycocalyx, which loses its clotting properties and releases significant amounts of blood clotting factors. The fibrin clot formed under such conditions is characterized by resistance to fibrinolysis and locally blocks blood vessels with the systemic development of endogenous intoxication. Destabilized proteins of the latter form micro- and nano-sized aggregates with a significant content of β-folded structures. This contributes to the increase of fibrin resistance to the proteolytic action of plasmin, causes the development of fibrosis of the tissues affected in this way, and leads to the failure of the functions of the relevant organs.

Selection of optimal conditions for obtaining malt from buckwheat

Buckwheat can be used in the production of specialized products (including therapeutic and preventive ones) for consumers suffering from gluten intolerance. Buckwheat malt in Russia is not produced on an industrial scale, it is purchased in European countries. It is necessary to know the parameters of processing domestic varieties of buckwheat for the production of buckwheat malt. Mathematical planning methods were used to predict the influence of the main factors of buckwheat germination on the activity of its hydrolytic enzymes for the production of malt with high enzymatic capacity. Dikul buckwheat was studied. Soaking of buckwheat was carried out by air-water method at a temperature of 15-16 °C in tap water to a relative humidity of 46-50 %, air and water pauses were alternated every 4 hours. Germination was carried out for 6 to 7 days at a temperature of 15 to 16 °C until the maximum activity of hydrolytic enzymes of amylolytic and proteolytic action was achieved. The sprouted grain was subjected to fermentation, for which it was left alone for 12-24 hours until the temperature in the middle layer rose to 50-55 °C, stirred and blown with conditioned air with such intensity that the humidity of the malt was 50-52 %, and the temperature in all layers was 67-68 °C for 5 days. Drying was carried out with a gradual increase in temperature from 67-68 °C to 74-75 °C for 16 hours to a humidity of 5-6 %. We predicted how the main factors of buckwheat germination affect the activity of its hydrolytic enzymes. The main factors affecting the activity of enzymes during germination: malting temperature, °C; malting humidity, %; malting duration, day. The criteria for evaluating the influence of the selected parameters were the abilities, units/g of dry substances: amylolytic, saccharifying, and proteolytic. For statistical processing of the study data, central rotatable uniform planning was used, which made it possible to obtain a regression equation that adequately describes the process of buckwheat germination in the course of 20 experiments in 3-fold repetition. To assess the adequacy of the mathematical model, an analysis of variance (ANOVA) of the experiment was performed in the Design Expert program (Stat-EaseInc.). The following values of the output parameters were obtained, units / g of dry substances – amylolytic capacity of 326.7, saccharifying capacity of 40.9 and proteolytic capacity of 381.9 units/g. at the appropriate input: temperature 15 °C; humidity 48 %; duration of malting 6 days.

Enzymatic transformation of wheat-flaxseed bran

Показана эффективность действия ферментных препаратов (ФП) целлюлолитического и протеолитического действия отечественного и зарубежного производства и мультиэнзимных композиций (МЭК) на их основе на биополимеры пшенично-льняных отрубей. The efficiency of the action of enzyme preparations (FP) of cellulolytic and proteolytic action of domestic and foreign production and multi-enzyme compositions (MEС) based on them on biopolymers of wheat-flax bran.

The protease MBTPS2 is an important regulator of several cellular processes, especially in health and sickness

Since the identification of MBTPS2 in 1997, tremendous progress has been made in determining the protease's functions. The protease has developed from an element of the SREBP cleavage machinery to an important regulator of several cellular processes, especially in health and sickness. With this newfound information from biochemical and structural biology, S2P's proteolytic action through peptide bond hydrolysis can occur in the membrane, providing a conceptual framework for appreciating S2P's roles in other aspects, and showing that many other substrates rely on S2P for their survival. In addition, we discovered the identity of both of S2P's catalytic active sites, an essential finding as the activity of the proteolysis as well as the pathogenesis of MBTPS2-caused illnesses seems to be connected to the molecular and biochemical features of the catalytic sites. Additionally, MBTPS2 causes different diseases, possibly illustrating the pleiotropic nature of the protein. Also, while the ailments reported thus far are all due to mutations that cause MBTPS2 to lose function, other variants that cause MBTPS2 to be hyperactive have not been examined. Nevertheless, recognizing the related sickness pathomechanism is a challenge. Pursuing these challenging technical areas would most definitely enhance our understanding of MBTPS2 in disease states. MBTPS2 appears to be nearing the solution to many of the remaining fundamental questions surrounding the mechanism of its action, as well as being a therapeutic target for new therapies.

Towards the Enhancement of Essential Oil Components’ Antimicrobial Activity Using New Zein Protein-Gated Mesoporous Silica Microdevices

The development of new food preservatives is essential to prevent foodborne outbreaks or food spoilage due to microbial growth, enzymatic activity or oxidation. Furthermore, new compounds that substitute the commonly used synthetic food preservatives are needed to stifle the rising problem of microbial resistance. In this scenario, we report herein, as far as we know, for the first time the use of the zein protein as a gating moiety and its application for the controlled release of essential oil components (EOCs). The design of microdevices consist of mesoporous silica particles loaded with essential oils components (thymol, carvacrol and cinnamaldehyde) and functionalized with the zein (prolamin) protein found in corn as a molecular gate. The zein protein grafted on the synthesized microdevices is degraded by the proteolytic action of bacterial enzymatic secretions with the consequent release of the loaded essential oil components efficiently inhibiting bacterial growth. The results allow us to conclude that the new microdevice presented here loaded with the essential oil component cinnamaldehyde improved the antimicrobial properties of the free compound by decreasing volatility and increasing local concentration.

Proteolytic enzyme arbitrated antagonization of helminthiasis by Cinnamomum cappara leaf extract in Pheretima posthuma

Background There have been several studies carried out to irradiate Helminthiasis however very little research have been carried out where in the enzymatic activity of plants are exploited to antagonize infections. Here we are analyzing the anthelmintic activity of Cinnamomum cappara leaf extract against Pheretima posthuma complimented by proteolytic action. Results The fresh leaves of Cinnamomum cappara was collected from local areas of Thrissur during December 2019. Plants were identified and authenticated by morphological and molecular characterization. The enzymatic action was analyzed by plotting Lineweaver–Burk plot which suggested that the extract possess the Km 185.77 μM for casein as substrate and obeyed Michaelis–Menten kinetics with typical hyperbolic relation with enzyme and increasing concentration of substrate. The effect of extract upon study subject was in directly proportional with concentration of antagonist where higher activities were obtained in high concentrations. The anatomical and histological studies suggested that the activity of extract was due to the degradation of muscular bundle of subject that resulted in the leakage of ceolomic fluid. Conclusions Cinnamomum cappara leaf extract possessed high degree of protease intervened anthelmintic activity against Pheretima posthuma. As the study subject show immense morphological and physiological resemblance with all other helminthic parasites, this results shall be adopted to further clinical and pharmacological applications.

Excitotoxicity and genetics of amyotrophic lateral sclerosis: effects of intracellular calcium accumulation on proteins encoded by the major genes underlying the disease

The aetiology of Amyotrophic Lateral Sclerosis (ALS), a fatal and incurable disease caused by motor neuron degeneration, is still poorly understood. The discovery of genetic forms of ALS helped to shed light on the mechanisms underlying this pathology, but also showed how complex these mechanisms are. Excitotoxicity is one of the processes strongly suspected to play a role in motor neuron degeneration in ALS. This process consists in neuron damage due to excessive intake of calcium ions (Ca2+) by the cell. This study aims to find a relationship between the proteins coded by the most relevant genes associated with ALS and excitotoxicity. In detail, the profile of eight proteins (TDP-43, C9ORF72, p62/SQSTM1, matrin3, VCP, FUS, SOD1 and profilin-1), was analysed in three different cell types induced to raise their cytoplasmic amount of Ca2+. Intracellular Ca2+ accumulation causes a significant decrease in the levels of TDP-43, C9ORF72, matrin3, VCP, FUS, SOD1 and profilin-1 and an increase in p62/SQSTM1. These events are associated to the proteolytic action of two proteases, calpains and caspases, as well as to the activation of autophagy, a process responsible for the degradation and recycling of cytoplasmic components. Interestingly, Ca2+ appears to both favour and hinder autophagy. The discovery of when Ca2+ levels become toxic for the cell, as well as understanding why the physiological processes of calpain proteolysis and autophagy become pathological, may elucidate the mechanisms responsible for ALS and help discover new therapeutic targets.

Identification of a Novel Thermostable Alkaline Protease from Bacillus megaterium-TK1 for the Detergent and Leather Industry

An increased need by the green industry for enzymes that can be exploited for eco-friendly industrial applications led us to isolate and identify a unique protease obtained from a proteolytic Bacillus megaterium-TK1 strain from a seawater source. The extracellular thermostable serine protease was processed by multiple chromatography steps. The isolated protease displayed a relative molecular weight (MW) of 33 kDa (confirmed by zymography), optimal enzyme performance at pH 8.0, and maximum enzyme performance at 70 °C with 100% substrate specificity towards casein. The proteolytic action was blocked by phenylmethylsulfonyl fluoride (PMSF), a serine hydrolase inactivator. Protease performance was augmented by several bivalent metal cations. The protease tolerance was studied under stringent conditions with different industrial dispersants and found to be stable with Surf Excel, Tide, or Rin detergents. Moreover, this protease could clean blood-stained fabrics and showed dehairing activity for cow skin with significantly reduced pollution loads. Our results suggest that this serine protease is a promising additive for various eco-friendly usages in both the detergent and leather industries.