Transpeptidase Activity Research Articles

Hematological and Serum Biochemistry Values in Free-Ranging Crested Porcupine.

The crested porcupine is a widespread naturalized Italian rodent of African origin; nevertheless, very little information on the population abundance and its health status is available. In this study, the hematological and serum chemistry profile of 10 free-ranging captured crested porcupines was established for the first time. The mean hematological values resulted: 5.7 SD 0.4 M/μL for red blood cells; 13.6 SD 0.8 g/dL for hemoglobin; 77.3 SD 5.7 fL for mean corpuscular volume and 30.1 SD 4.7 g/dL for mean corpuscular hemoglobin concentration; 14.4 SD 7.2 K/μL for white blood cells; and 557.0 SD 469.9 K/μL for platelets. The mean urea and creatinine values resulted with 19.8 SD 8.3 mg/dL and 1.6 SD 3.0 mg/dL, respectively. The mean value of total protein was 6.7 SD 1.0 g/dL, with values of albumin higher than globulins. The mean activity of creatine kinase, aspartate transaminase, gamma-glutamyl transpeptidase, and alkaline phosphatase was 927.3 SD 607.6 U/L, 199.2 SD 70.8 U/L, 16.9 SD 13.7 U/L, and 256 SD 75.8 U/L, respectively. Highest values of alkaline phosphatase were recorded in two porcupines presenting severe injuries with clear signs of infection. These preliminary results may be a helpful tool in order to assess porcupine health status.

In Vivo Visualization of γ-Glutamyl Transpeptidase Activity with an Activatable Self-Immobilizing Near-Infrared Probe.

γ-Glutamyl transpeptidase (GGT), a type of cell membrane-bound enzyme, is closely involved in a wide range of physiological and pathological processes, and a large number of fluorogenic probes have been developed to detect the activity of GGT. However, the use of these imaging reagents to visualize GGT activity in vivo is largely limited because of rapid diffusion and clearance of activated fluorophores. Herein, by merging quinone methide and a fluorogenic enzyme substrate, we report an activatable self-immobilizing near-infrared probe for the in vitro and in vivo imaging of GGT activity. This probe is initially fluorescently silent, but the selective activation by GGT is able to significantly increase its fluorescence intensity at 714 nm and covalently anchor activated fluorophores at the site of interest. We have shown that this probe induced a much stronger fluorescence on live GGT-overexpressing cells compared to regular fluorogenic probes and allowed wash-free and real-time imaging of enzyme activity. More importantly, the use of this probe in the imaging of GGT activity in U87MG tumor-bearing mice by i.v. administration indicates that this self-immobilizing reagent is capable of efficiently enhancing its retention at the detection target and thus leads to much improved detection sensitivity compared to regular fluorogenic probes. This study demonstrates the advantage of fluorogenic probes with activatable anchors in the noninvasive imaging of enzyme activity in highly dynamic in vivo systems.

Designed Streptococcus pyogenes Sortase A Accepts Branched Amines as Nucleophiles in Sortagging.

Sortase-mediated ligation (sortagging) is commonly performed using the Staphylococcus aureus sortase A (SaSrtA) that strictly recognizes the N-terminal glycine residue. In this work, a rational design of Streptococcus pyogenes sortase A (SpSrtA) for improved transpeptidase activity toward different N-terminal amino acid residues was conducted. The generated variant SpSrtA M3 (E189H/V206I/E215A) showed up to 6.6-fold (vs SpSrtA wild-type) enhanced catalytic efficiency. Additionally, M3 retains the specificity toward N-terminal alanine, glycine, serine residues, as well as branched (at α-carbon) primary amines as wild-type parent. Furthermore, M3 was applied for head-to-tail backbone cyclization of proteins.

Novel Perfluoroalkyl Ether Carboxylic Acids (PFECAs) and Sulfonic Acids (PFESAs): Occurrence and Association with Serum Biochemical Parameters in Residents Living Near a Fluorochemical Plant in China.

Although perfluoroalkyl ether carboxylic (PFECAs) and sulfonic acids (PFESAs) have been widely detected in environmental matrices, their occurrence in humans and impact on human health remains insufficiently understood. Here, we report on 13 PFECAs and PFESAs in 977 sera samples collected from residents living near a fluorochemical plant in Shandong, China. The sum concentration of these emerging PFECAs accounted for 13% of the total PFASs in the serum of the participants, with the frequent detection of several PFECAs (>95%) (PFMOAA, PFO4DA, and PFO5DoDA at median concentrations of 12.91, 0.142, and 0.987 ng/mL, respectively) and PFESAs (98.7%) (Nafion byproduct 2 at a median concentration of 0.097 ng/mL). Serum PFMOAA, PFO5DoDA, and 6:2 Cl-PFESA levels were significantly higher in males than in females. Positive relationships were observed between age and PFMOAA, 6:2 Cl-PFESA, and H-PFMO2OSA levels, whereas HFPO-TA and PFO5DoDA serum concentrations in the 0-40-year age group were lower than that in the >40-year age group. Furthermore, multivariate linear regression models and sensitivity analyses showed positive associations among PFO5DoDA levels, elevated lipid parameters (cholesterol, low-density lipoprotein cholesterol, and triglycerides), liver function markers (albumin levels and alanine transaminase, aspartate aminotransferase, and glutamyl transpeptidase activities), and uric acid levels. Thus, our results suggest potential health risks from exposure to novel PFESAs and PFECAs (especially PFO5DoDA).

Structural and functional studies of Arabidopsis thaliana legumain beta reveal isoform specific mechanisms of activation and substrate recognition

The vacuolar cysteine protease legumain plays important functions in seed maturation and plant programmed cell death. Because of their dual protease and ligase activity, plant legumains have become of particular biotechnological interest, e.g. for the synthesis of cyclic peptides for drug design or for protein engineering. However, the molecular mechanisms behind their dual protease and ligase activities are still poorly understood, limiting their applications. Here, we present the crystal structure of Arabidopsis thaliana legumain isoform β (AtLEGβ) in its zymogen state. Combining structural and biochemical experiments, we show for the first time that plant legumains encode distinct, isoform-specific activation mechanisms. Whereas the autocatalytic activation of isoform γ (AtLEGγ) is controlled by the latency-conferring dimer state, the activation of the monomeric AtLEGβ is concentration independent. Additionally, in AtLEGβ the plant-characteristic two-chain intermediate state is stabilized by hydrophobic rather than ionic interactions, as in AtLEGγ, resulting in significantly different pH stability profiles. The crystal structure of AtLEGβ revealed unrestricted nonprime substrate binding pockets, consistent with the broad substrate specificity, as determined by degradomic assays. Further to its protease activity, we show that AtLEGβ exhibits a true peptide ligase activity. Whereas cleavage-dependent transpeptidase activity has been reported for other plant legumains, AtLEGβ is the first example of a plant legumain capable of linking free termini. The discovery of these isoform-specific differences will allow us to identify and rationally design efficient ligases with application in biotechnology and drug development.

Oridonin induces ferroptosis by inhibiting gamma-glutamyl cycle in TE1 cells.

Oridonin (Ori) is a natural tetracyclic diterpenoid active compound with excellent antitumor activity, but the mechanism of Ori on esophageal cancer cell, TE1, remains unclear. In this study, we examined the levels of intracellular iron, malondialdehyde, and reactive oxygen species after Ori treatment, while interfering with the effects of Ori with ferroptosis inhibitor, demonstrating that Ori's inhibition of TE1 cell proliferation is associated with ferroptosis. To understand the molecular mechanism of Ori, we performed UPLC-MS/MS metabolomics profiling on TE1 cells, which show that gamma-glutamyl amino acids (gamma-glutamylleucine, gamma-glutamylvaline), 5-oxoproline, glutamate, GSH, and GSSG are changed significantly after Ori treatment. Meanwhile, the activity of gamma-glutamyl transpeptidase 1 (GGT1) decreased. This revealed that Ori inhibited the gamma-glutamyl cycle in TE1 cells. Furthermore, we found that Ori can covalently bind to cysteine to form the conjugate oridonin-cysteine (Ori-Cys), resulting in the inhibition of glutathione synthesis, which is consistent with the decrease in the enzymatic activity of glutamate cysteine ligase catalytic subunit (GCLC). Eventually, the value of intracellular GSH/GSSG was reduced, and the enzymatic activity of the glutathione peroxidase 4 (GPX4) was significantly decreased. In conclusion, our experiments indicated that Ori can inhibit the gamma-glutamyl cycle, thereby inducing ferroptosis to exert anti-cancer activity.

Body Fat Changes and Liver Safety in Obese and Overweight Women Supplemented with Conjugated Linoleic Acid: A 12-Week Randomised, Double-Blind, Placebo-Controlled Trial.

Preliminary evidence suggests that conjugated linoleic acid (CLA) may reduce body weight and affect body composition. The present study assessed the effect of CLA supplementation on body fat composition in overweight and obese women, while also evaluating the liver safety of CLA use. Seventy-four obese or overweight women were randomly assigned to receive 3 g/day CLA or placebo for 12 weeks. Body composition (dual-energy X-ray absorptiometry) and liver function (13C-methacetin breath test and serum liver enzymes) were assessed before and after the trial. Patients receiving CLA experienced a significant reduction of total body fat expressed as mass (p = 0.0007) and percentage (p = 0.0006), android adipose tissue (p = 0.0002), gynoid adipose tissue (p = 0.0028), and visceral adipose tissue (p = 4.2 × 10−9) as well as a significant increase in lean body mass to height (p = 6.1 × 10−11) when compared to those receiving a placebo. The maximum momentary 13C recovery changes and end-point values were significantly higher in the CLA group when compared to the placebo group (p = 0.0385 and p = 0.0076, respectively). There were no significant changes in alanine aminotransferase, asparagine aminotransferase, and gamma-glutamyl transpeptidase activities between the groups. In conclusion, CLA supplementation was well tolerated and safe for the liver, which shows beneficial effects on fat composition in overweight and obese women.

Enzymodiagnostics of ophthalmoherpes complications

The purpose of the studyis to evaluate the possibility of using enzyme analysis of the tear and oral fluids to diagnose the complications of ophthalmoherpes in children. In case of herpetic keratitis in children, the distinctive features of enzyme activity are revealed depending on the clinical form and severity of ophthalmoherpes. This allows you to assess the intensity of inflammation, the effectiveness of treatment and predict the complications of the disease. Determination of γ-glutamyl transpeptidase activity in tears can be used to identify myopia in children of preschool age. The study of the tear fluid allows you to expand the possibilities of non-invasive diagnosis of ophthalmoherpes in children.

Acute exposure to bis(2-ethylhexyl)phthalate disrupts calcium homeostasis, energy metabolism and induces oxidative stress in the testis of Danio rerio

Bis(2-ethylhexyl)phthalate (BEHP) negatively affects testicular functions in different animal species, disturbing reproductive physiology and male fertility. The present study investigated the in vitro acute effect of BEHP on the mechanism of action of ionic calcium (Ca2+) homeostasis and energy metabolism. In addition, the effect of BEHP on oxidative stress was studied in vitro and in vivo in the testis of Danio rerio (D. rerio). Testes were treated in vitro for 30 min with 1 μM BEHP for 45Ca2+ influx measurements. Testes were also incubated with 1 μM BEHP for 1 h (in vitro) or 12 h (in vivo) for the measurements of lactate content, 14C-deoxy-d-glucose uptake, lactate dehydrogenase (LDH) and gamma-glutamyl transpeptidase (GGT) activity, total reactive oxygen species (ROS) production and lipid peroxidation. In addition, the effect of BEHP (1 μM) on GGT, glutamic oxaloacetic transferase (GOT) and glutamic pyruvic transferase (GPT) activity in the liver was evaluated after in vivo treatment for 12 h. BEHP disturbs the Ca2+ balance in the testis when given acutely in vitro. BEHP stimulated Ca2+ influx occurs through L-type voltage-dependent Ca2+ channels (L-VDCC), transitory receptor potential vaniloid (TRPV1) channels, reverse-mode Na+/Ca2+ exchanger (NCX) activation and inhibition of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA). BEHP affected energy metabolism in the testis by decreasing the lactate content and LDH activity. In vitro and in vivo acute effects of BEHP promoted oxidative stress by increasing ROS production, lipid peroxidation and GGT activity in the testis. Additionally, BEHP caused liver damage by increasing GPT activity.

Structure of the Peptidoglycan Synthase Activator LpoP in Pseudomonas aeruginosa

SummaryPeptidoglycan (PG) is an essential component of the bacterial cell wall and is assembled from a lipid II precursor by glycosyltransferase and transpeptidase reactions catalyzed in particular by bifunctional class A penicillin-binding proteins (aPBPs). In the major clinical pathogen Pseudomonas aeruginosa, PBP1B is anchored within the cytoplasmic membrane but regulated by a bespoke outer membrane-localized lipoprotein known as LpoP. Here, we report the structure of LpoP, showing an extended N-terminal, flexible tether followed by a well-ordered C-terminal tandem-tetratricopeptide repeat domain. We show that LpoP stimulates both PBP1B transpeptidase and glycosyltransferase activities in vitro and interacts directly via its C terminus globular domain with the central UB2H domain of PBP1B. Contrary to the situation in E. coli, P. aeruginosa CpoB does not regulate PBP1B/LpoP in vitro. We propose a mechanism that helps to underscore similarities and differences in class A PBP activation across Gram-negative bacteria.

Class A PBPs have a distinct and unique role in the construction of the pneumococcal cell wall

In oval-shaped Streptococcus pneumoniae, septal and longitudinal peptidoglycan syntheses are performed by independent functional complexes: the divisome and the elongasome. Penicillin-binding proteins (PBPs) were long considered the key peptidoglycan-synthesizing enzymes in these complexes. Among these were the bifunctional class A PBPs, which are both glycosyltransferases and transpeptidases, and monofunctional class B PBPs with only transpeptidase activity. Recently, however, it was established that the monofunctional class B PBPs work together with transmembrane glycosyltransferases (FtsW and RodA) from the shape, elongation, division, and sporulation (SEDS) family to make up the core peptidoglycan-synthesizing machineries within the pneumococcal divisome (FtsW/PBP2x) and elongasome (RodA/PBP2b). The function of class A PBPs is therefore now an open question. Here we utilize the peptidoglycan hydrolase CbpD that targets the septum of S. pneumoniae cells to show that class A PBPs have an autonomous role during pneumococcal cell wall synthesis. Using assays to specifically inhibit the function of PBP2x and FtsW, we demonstrate that CbpD attacks nascent peptidoglycan synthesized by the divisome. Notably, class A PBPs could process this nascent peptidoglycan from a CbpD-sensitive to a CbpD-resistant form. The class A PBP-mediated processing was independent of divisome and elongasome activities. Class A PBPs thus constitute an autonomous functional entity which processes recently formed peptidoglycan synthesized by FtsW/PBP2×. Our results support a model in which mature pneumococcal peptidoglycan is synthesized by three functional entities, the divisome, the elongasome, and bifunctional PBPs. The latter modify existing peptidoglycan but are probably not involved in primary peptidoglycan synthesis.

Determination of the activity of γ-glutamyl transpeptidase and of its inhibitors by using the inner filter effect on the fluorescence of nitrogen-doped carbon dots.

A fluorescence (FL) probe for determination of γ-glutamyl transpeptidase (GGT) activity and evaluation of inhibitors was developed based on the inner filter effect (IFE) of nitrogen-doped carbon dots (N-CDs). Bright green emissive N-CDs were synthesized by one-step hydrothermal technique with catechol and ethylenediamine. The excitation and emission wavelengths for N-CDs were 408 and 510nm, respectively. γ-L-Glutamyl-4-nitroanilide (γ-G4NA) was employed as the substrate of GGT. The absorption spectrum of GGT catalytic product (4-nitroaniline, 4-NA) overlapped greatly with the excitation spectrum of N-CDs. 4-NA acted as the absorber in IFE to quench the FL of N-CDs. Thus, the FL quenching of N-CDs was closely related to GGT activity. The established FL method offered good linear relationship within 2.0-10.0UL-1 (R2, 0.982) and 10.0-110.0UL-1 (R2, 0.998) with a low detection limit of 0.6UL-1. The method was successfully applied to investigate GGT activity in human serum samples with acceptable recoveries (99.1-105.0%). The approach was also employed for screening GGT inhibitors from different polar extracts of Schisandra chinensis. Results indicated that this strategy presents superior characteristics for GGT sensing. This method has great potential as a candidate for diagnosis of GGT-related diseases and high-throughput drug discovery. Graphical abstract.

PBP1a glycosyltransferase and transpeptidase activities are both required for maintaining cell morphology and envelope integrity in Shewanella oneidensis.

In rod-shaped Gram-negative bacteria, penicillin binding protein 1a (PBP1a) and 1b (PBP1b) form peptidoglycan-synthesizing complexes with the outer membrane lipoprotein LpoA and LpoB, respectively. Escherichia coli mutants lacking PBP1b/LpoB are sicker than those lacking PBP1a/LpoA. However, we previously found that mutants lacking PBP1a/LpoA but not PBP1b/LpoB are deleterious in Shewanella oneidensis. Here, we show that S. oneidensis PBP1a (SoPBP1a) contains conserved signature motifs with its E. coli counterpart, EcPBP1a. Although EcPBP1a play a less prominent role in E. coli, it is capable of substituting for the SoPBP1a in a manner dependent on SoLpoA. In S. oneidensis, expression of PBP1b is lower than PBP1a, and therefore the additional expression of SoPBP1b at low levels can functionally compensate for the absence of SoPBP1a. Importantly, S. oneidensis PBP1a variants lacking either glycosyltransferase (GTase) or transpeptidase (TPase) activity fail to maintain normal morphology and cell envelope integrity. Similarly, SoPBP1b variants also fail to compensate for the loss of SoPBP1a. Furthermore, overproduction of variants of SoPBP1a, but not SoPBP1b, has detrimental effects on cell morphology in S. oneidensis wild type cells. Overall, our results indicate that the combined enzymatic activities of SoPBP1a are essential for cell wall homeostasis.

Partially shielded enzymes capable of processing large protein substrates.

We report the first method of enzyme protection enabling the production of partially shielded enzymes capable of processing substrates as large as proteins. We show that partially shielded sortase retains its transpeptidase activity and can perform bioconjugation reactions on antibodies. Moreover, a partially shielded trypsin is shown to outperform its soluble counterpart in terms of proteolytic kinetics. Remarkably, partial enzyme shielding results in a drastic increase in temporal stability of the enzyme.

Effects of Bacillus cereus and coumarin on growth performance, blood biochemical parameters, and meat quality in broilers.

Background and Aim:Progressive antibiotic resistance has become the primary threat to public health. The search for alternative substances with similar effects is now a global challenge for poultry farming. The aim of this study was to investigate the action of the probiotic Bacillus cereus (BC) and coumarin (CO) on broiler productivity, biochemical indicators of blood, and muscular and liver tissues.Materials and Methods:The trial of this study included Arbor Acres cross broiler chickens that were grown up to the age of 42 days. The experiment was conducted on 200 broiler chickens divided into four experimental groups of 50 individuals each: The control group received ration without additives (main ration [MR]), the first experimental group received MR+BC, the second received MR+CO, and the third received −MR+BC+CO. A biochemical and hematological analyzer was used to estimate elemental concentrations using inductively coupled plasma mass spectrometry and inductively coupled plasma atomic emission spectrometry.Results:Inclusion of CO and CO+BC in the diet improved growth rates and reduced feed consumption (FC) per kg of live weight gain. Decreased white blood cell count, increased creatinine and triglycerides (CO), changes in aminotransferase and transpeptidase activity, and increases in chemical elements in the liver and pectoral muscles (BC+CO) were observed. The inclusion of BC+CO in the diet contributed to increases in a greater number of chemical elements in the liver (calcium [Ca], K, magnesium, Mn, Si, and Zn) and the pectoral muscles (Ca, Na, Co, Cu, Fe, Mn, Ni, and Zn).Conclusion:The inclusion of CO and CO+BC in the diet improves growth rates and reduces FC in broilers against a background of the absence of mortality during the experiment.

Sensitive and selective SERS probe for detecting the activity of γ-glutamyl transpeptidase in serum

γ-Glutamyl transpeptidase (GGT) has attracted considerable attention for its regulatory effect on glutathione metabolism in living organisms; further, its close relationship with physiological dysfunctions such as hepatitis and liver cancers has enhanced its applicability. Therefore, the accurate detection of GGT levels is particularly important for the early diagnosis of diseases. Thus, we herein report the development of a surface-enhanced Raman spectroscopic (SERS) probe, namely bis-s,sʹ-((s)-4,4ʹ-thiolphenylamide-Glu) (b-(s)-TPA-Glu), that comprises of a γ-glutamyl moiety for detection of the GGT activity. In this system, detection was achieved by observing differences in the SERS spectral profiles of the b-(s)-TPA-Glu probe and its corresponding hydrolysis product that resulted from the catalytic action of GGT. This SERS probe system exhibited a high selectivity toward GGT due to a combination of its specific catalytic action and the distinctive spectroscopic fingerprint of the SERS technique. The developed SERS approach was also found to be approximately linear in the range of 0.2–200 U/L, and a limit of detection of 0.09 U/L was determined. Furthermore, the proposed SERS method was suitable for detection of the GGT activity of clinical serum samples and also for evaluation of the inhibitors of GGT. Consequently, this approach is considered to be a promising diagnostic and drug screening tool for GGT-associated diseases.

Toxicological characteristics of the cinnamic

The toxic properties of the cinnamic alcohol with an aim of its hygienic rating are studied. DL for male rats, male and female mice are respectively 3300, 3000 and 2700 mg/kg. It refers to the substances of hazard class 3. The clinical picture of acute poisoning was characterized by the general depression, muscle relaxation, disorder of the movement coordination, decrease in body temperature and death in the first and the second days after the poisoning. The differences in sex and species sensitivity of the animals to the substance are not observed: the rates of species differences and sex sensitivity are ~1. Single and repeated (10 days) inhalation of the vapors of the cinnamic alcohol at the greatest possible saturation neither cause the death of mice no signs of toxicity. It has a weak ability to cumulation: the cumulation coefficient is 5.4. It has no local irritating action to the skin. Skin-resorptive and sensitizing effects are not revealed. It has a weak irritating effect on the mucous membranes of the eyes. In the subacute experiment after the introduction of the substance into the stomach in a total dose equal to 5.2 DL we noted the lag of the experimental animals in body weight gain from the control ones, the increase in the activity of y-glutamyl transpeptidase and transaminases in serum. In the peripheral blood the decrease in hemoglobin, erythrocytes and eosinophils count is revealed. The threshold of the acute inhalation effect is 140 mg/m (by the reduction in the number of erythrocytes and hemoglobin in peripheral blood). Tentative safe level of the exposure to the cinnamic alcohol in the air of the working zone is 5 mg/m (vapors).

Enzymatic synthesis of L-theanine from L-glutamine and ethylamine by Bacillus licheniformis γ-glutamyltranspeptidase and its mutants specialized in transpeptidase activity

Abstract In this investigation, Bacillus licheniformis γ-glutamyltranspeptidase (BlGGT) together with its two transpeptidase-specialized mutants (N450D and N450Q) were employed for the biocatalytic synthesis of L -theanine. A high-pressure liquid chromatographic method coupled with ultraviolet detection (HPLC/UV) was used to quantify L -theanine content in the reaction mixture. Through a series of experiments, the perfect conditions for the biocatalytic synthesis were found to be an operational pH of 10.5, a reaction time of 4 h, L -glutamine and ethanolamine in a 1:2.4 M ratio, and enzymes at a working concentration of 25 μg/mL. In a batch process for N450D-mediated synthesis of L -theanine at 37 °C, a conversion rate of ~94% was achieved from 250 mM l -glutamine and 600 mM ethylamine. Successful synthesis of the desired product was further verified by mass spectrometry analysis. Conclusively, the experimental results suggest a high potential application in the production of L -theanine by BlGGT-mediated biocatalysis.

Role Of Β-Carotene Against Toxic Effect Of Titanium Dioxide Nanoparticles On Cerebral Cortex Of Adult Albino Rat: Histological And Biochemical Approach

Objective: Detect the neurotoxic effect of titanium dioxide nanoparticles by investigation of the biochemical and histological changes in cerebral cortex of albino rats, and to evaluate the potential protective role of β-carotene against this toxicity.Methods: Titanium dioxide nanoparticles (TiO2 NPs) administered intraperitoneal 100, 300 mg/kg, daily for 14 days besides one protected group received 10mg/kg β-carotene by gavage for 7 days prior and 14 days with 300 mg/kg nanoparticles administration, along with one control group and one group received 10mg/kg β-carotene by gavage for 21 days. Histological and histochemichal cerebral cortical tissue examination and standard biochemical methods for estimation of enzymes superoxide dismutase, glutathione peroxidase, catalase, alkaline phosphatase, gamma-glutamyl transpeptidase, and 5’ nucleotidase were implemented.Results: In contrast to administration of low dose TiO2 NPs, administration of high dose TiO2 NPs led to marked histological changes in form of decrease number of vesicular neurons and loss of background homogeneity with appearance of vacuolations and hydropic changes. Moreover, increase in astrocytes number with appearance of area of gliosis and increase in apoptosis index. Biochemical analysis showed significant decreases in the activities of the antioxidant enzymes: superoxide dismutase, glutathione peroxidase and catalase. Besides, significant decrease in the activities of alkaline phosphatase and gamma-glutamyl transpeptidase, although the activity of 5’-nucleotidase was not inclined significantly by administration of nanoparticles. These changes didn’t markedly appear with the use of β-carotene.Conclusion: High dose TiO2 NPs has neurotoxic effects but application of β-carotene may serve as a beneficial medication to protect against neurotoxicity induced by nanoparticles.

Sulfur-containing histidine compounds inhibit γ-glutamyl transpeptidase activity in human cancer cells

γ-Glutamyl transpeptidase (GGT) is an enzyme located on the surface of cellular membranes and involved in GSH metabolism and maintenance of redox homeostasis. High GGT expression on tumor cells is associated with increased cell proliferation and resistance against chemotherapy. GGT inhibitors evaluated so far in clinical trials are too toxic for human use. In this study, using enzyme kinetics analyses, we demonstrate that ovothiols, 5(Nπ)-methyl thiohistidines of marine origin, act as noncompetitive inhibitors of GGT, with an apparent Ki of 21 μm, when we fixed the concentrations of the donor substrate. We found that these compounds are more potent than the known GGT inhibitor 6-diazo-5-oxo-l-norleucine and are not toxic toward human embryonic cells. In particular, cellular process-specific fluorescence-based assays revealed that ovothiols induce a mixed cell-death phenotype of apoptosis and autophagy in GGT-overexpressing cell lines, including human liver cancer and chronic B leukemic cells. The findings of our study provide the basis for further development of 5-thiohistidines as therapeutics for GGT-positive tumors and highlight that GGT inhibition is involved in autophagy.