Prediction of the three-dimensional structure of aflatoxin of Aspergillus flavus by homology modelling

Aflatoxins are polyketide-derived secondary metabolites produced by Aspergillus spp. The toxic effects of aflatoxins have adverse consequences for human health and agricultural economics. The aflR gene, a regulatory gene for aflatoxin biosynthesis, encodes a protein containing a zinc-finger DNA-binding motif. AFLR-Protein three-dimensional model was generated using Robetta server. The modeled AFLR-Protein was further optimization and validation using Rampage. In the simulations, we monitored the backbone atoms and the C-α-helix of the modeled protein. The low RMSD and the simulation time indicate that, as expected, the 3D structural model of AFLR-protein represents a stable folding conformation. This study paves the way for generating computer molecular models for proteins whose crystal structures are not available and which would aid in detailed molecular mechanism of inhibition of aflatoxin.     

Effect of training on treadmill performance,aerobic capacity,and body responses to acute cold exposure

An attempt was made to test the hypothesis that regular physical activity at the anaerobic threshold can stimulate an increase in the amount of brown or beige body fat, which can manifest itself in increased lactate utilization during exercise and increased reactivity in response to acute regional cooling. The methods used in the study included the ramp test; regional acute cold exposure; measurement of gas exchange; lactate and glucose in the blood; heart rate; heart rate, blood pressure, and respiration variability at rest and during standard functional tests; infrared thermal imaging; and statistical methods of analysis of results. Training of ten physically active volunteers (7 males and 3 females) on a treadmill at a speed corresponding to 75–80% of personal maximum oxygen consumption \(\left( {V_{O_{2\max } } } \right)\) for 30 min 3 times per week at a fixed ambient temperature of 21–22°C for 6 weeks resulted in a significant (from 19 to 39%) increase in exercise duration in the ramp test, whereas \(V_{O_{2\max } }\) changed, on average, only slightly. The increase in the anaerobic threshold power was associated with a sharp slowdown in the accumulation of lactate during the ramp test. The lactate utilization rate during the recovery period, on the contrary, increased. In general, work efficiency during test load significantly increased. Noticeable changes in the condition and responses to the standard functional tests of the autonomic system were not found, as judged by the heart rate variability, blood pressure, and respiration variability at rest and during orthostatic tests and imposed breathing rhythm. The functional response of the body to acute cold exposure (1-min cooling of the feet in ice water) did not change after a cycle of training, both in terms of metabolism (oxygen consumption, etc.) and the skin temperature dynamics in the areas of most probable location of brown adipose tissue (BAT). These data do not confirm our previous hypothesis (2010) about the function of BAT as a universal homeostatic instrument in the body. Probably, if the formation of the beige adipose tissue is stimulated by physical activity and hormone irisin, produced by muscles, this tissue is involved in lactate utilization but is not involved in the thermoregulatory responses.     

Mapping the amino acid properties of constituent nucleoporins onto the yeast nuclear pore complex

Visualization of molecular structures aids in the understanding of structural and functional roles of biological macromolecules. Macromolecular transport between the cell nucleus and cytoplasm is facilitated by the nuclear pore complex (NPC). The ring structure of the NPC is large and contains several distinct proteins (nucleoporins) which function as a selective gate for the passage of certain molecules into and out of the nucleus. In this note we demonstrate the utility of a python code that allows direct mapping of the physiochemical properties of the constituent nucleoporins on the scaffold of the yeast NPC׳s cytoplasmic view. We expect this tool to be useful for researchers to visualize the NPC based on their physiochemical properties and how it alters when specific mutations are introduced in one or more of the nucleoporins. The code developed using Python is available freely from the authors.     

The composition, structural properties and binding of very-low-density and low-density lipoproteins to the LDL receptor in normo- and hypertriglyceridemia: relation to the apolipoprotein E phenotype

The composition, apolipoprotein structure and lipoprotein binding to the LDL receptor were studied for very-low-density (VLDL) and low-density lipoprotein (LDL) particles isolated from subjects with apoE phenotype E3/3 (E3), E2/2 or E2/3 (E2+) and E3/4 or E4/4 (E4+) and a wide range of plasma triglyceride (TG) contents. The data combined for all three phenotype groups can be summarized as follows. (i) A decrease in accessibility of VLDL tryptophan residues to I- anions with a decrease in tryptophan surface density, concomitant with an increase in VLDL dimensions, reflects the increased efficiency of protein-protein interactions. (ii) A gradual increase in the quenching constant for LDL apoB fluorescence with an increase in TG/cholesterol (Chol) ratio reflects the 'freezing' effect of Chol molecules on apoB dynamics. (iii) Different mechanisms specific for a particular lipoprotein from E3/3 or E2/3 subjects are responsible for apoE-mediated VLDL binding and apoB-mediated LDL binding to the LDL receptor in a solid-phase binding assay. (iv) The 'spacing' effect of apoC-III molecules on apoE-mediated VLDL binding results in a decrease in the number of binding sites. (v) The maximum of the dependence of the LDL binding affinity constant on relative tryptophan density corresponds to LDL intermediate size. VLDL particles from hypertriglyceridemic E2/3 heterozygotic individuals had remnant-like properties (increased cholesterol, apoE and decreased apoC-III content) while their binding efficiency was unchanged. Based on the affinity constant value and LDL-Chol content, increased competition between VLDL and LDL for the binding to the LDL receptor upon increase in plasma TG is suggested, and LDL from hypertriglyceridemic E3/3 homozygotic individuals is the most efficient competitor.     

Method to Estimate Activity of Subcutaneous Thermogenic Structures on Exposure to Stimuli of Different Modalities

A method to estimate activity of subcutaneous thermogenic structures was developed on the basis of dynamical infrared thermography. The gist is comparing the dynamics of skin temperature changes for different body sites on exposure to factors evoking metabolic responses. The method can be used for investigating the mechanisms of changes in body temperature in response to various disturbing factors and, in certain cases, can help to differentiate metabolic and vascular responses.     

Structural peculiarities of the binding of very low density lipoproteins and low density lipoproteins to the LDL receptor in hypertriglyceridemia: role of apolipoprotein E

Very low (VLDL) and low density lipoproteins (LDL) were isolated from plasma of patients with the E3/3 phenotype which were divided into three groups based on their plasma triglyceride content: low (TG<200 mg/dl, TG(l)), intermediate (200<300 mg/dl, TG(i)300 mg/dl, TG(h)). The protein density (PD) on the VLDL and LDL surface was calculated from lipoprotein composition and protein location was studied by tryptophan fluorescence quenching by I(-) anions at 25 degrees C and 40 degrees C. A comparison of the TG(h) with the TG(l) group revealed a significant (<0.05) increase of the PD parameter as much as 21% for VLDL, but not for LDL where this parameter did not change for any group; generally, PD(LDL) values were 3.2-3.8-fold lower than PD(VLDL). In accordance with this difference, the tryptophan accessibility f in VLDL vs. LDL was lower at both temperatures. There were temperature-induced changes of the f parameter in opposite directions for these lipoproteins. The difference in f value gradually decreased for VLDL in the direction TG(l)TG(i)TG(h) while for LDL there was a U-shaped dependence for these groups. The Stern-Volmer quenching constant K(S-V) which is sensitive to both temperature and viscosity, did not change for VLDL, but K(S-V)(LDL) was 2-3-fold higher for the TG(i) group compared to the other two. The efficiencies of VLDL and LDL binding to the LDL receptor (LDLr) in vitro were compared by solid-phase assay free of steric hindrance observed in cell binding. The maximal number of binding sites did not change for either type of particles and between groups. The association constant K(a) and apolipoprotein (apo) E/apoB mole ratio values all increased significantly for VLDL, but not for LDL, in comparison of the TG(i+h) with the TG(l) group. Based on VLDL and LDL concentrations in serum and on the affinity constant values obtained in an in vitro assay, VLDL concentrations corresponding to 50% inhibition of LDL binding (IC(50)) were calculated in an assumption of the competition of both ligands for LDLr in vivo; the mean values of IC(50) decreased 2-fold when plasma TG exceeded 200 mg/dl. The functional dependences of K(a)(VLDL), IC(50) and apoE content in VLDL (both fractional and absolute) and in serum on TG content in the whole concentration range studied were fitted to a saturation model. For all five parameters, the mean half-maximum values TG(1/2) were in the range 52-103 mg/dl. The efficiency of protein-protein interactions is suggested to differ in normolipidemic vs. HTG-VLDL and apoE content and/or protein density on VLDL surface may be the primary determinant(s) of the increased binding of HTG-VLDL to the LDL receptor. ApoCs may compete with apoE for the binding to the VLDL lipid surface as plasma triglyceride content increases. The possible competition of VLDL with LDL for the catabolism site(s) in vivo, when plasma TG increases, could explain the atherogenic action of TG-rich lipoproteins. Moreover, the 'dual action' hypothesis on anti-atherogenic action of apoE-containing high density lipoproteins (HDL) in vivo is suggested: besides the well-known effect of HDL as cholesteryl ester catabolic outway, the formation of a transient complex of apoE-containing discs appearing at the site of VLDL TG hydrolysis by lipoprotein lipase with VLDL particles proposed in our preceding paper promotes the efficient uptake of TG-rich particles; in hypertriglyceridemia due to the diminished HDL content this uptake seems to be impaired which results in the increased accumulation of the remnants of TG-rich particles. This explains the observed increase in cholesterol and triglyceride content in VLDL and LDL, respectively, due to the CETP-mediated exchange of cholesteryl ester and triglyceride molecules between these particles.     

The treatment of isolated genera

Problems presented by genera, or small groups of genera, which have been given family rank are reviewed, and the genera are divided into a number of categories according to the closeness of their affinity to other genera or families. Satellite genera that stand in close relation to families should be united with them. Binary families, that have been divided into two (or more) related families, should be re–united. Families connected by linking genera, should, logically, be united but practical considerations usually prevent this. Clusters of diverse but more or less distantly related genera present unusual problems, being treated either as several, often monogeneric families or as a loosely structured family. Truly isolated genera must be given family and often ordinal rank.     

Activity of tissue factor in microparticles produced <Emphasis Type="Italic">in vitro</Emphasis> by endothelial cells,monocytes, granulocytes,and platelets

Activity of tissue factor (TF) in membrane microparticles (MPs) produced in vitro by endothelial cells (ECs), monocytes, THP-1 monocytic cells, granulocytes, and platelets was investigated. ECs were isolated from human umbilical vein, and monocytes, granulocytes, and platelets–from the blood of healthy donors. ECs, monocytes, and THP-1 cells were activated by bacterial lipopolysaccharide, granulocytes–by lipopolysaccharide or phorbol myristate acetate, and platelets - by SFLLRN, thrombin receptor-activating peptide. MPs were sedimented from the culture medium or supernatant of activated cells at 20,000g for 30 min. Coagulation activity of MPs was analyzed in a modified recalcification assay by assessing their effects on coagulation of donor plasma depleted of endogenous MPs (by centrifuging at 20,000g for 90 min). MPs from all cell types accelerated plasma coagulation. Antibodies blocking TF activity prolonged coagulation lagphase in the presence of MPs from ECs, monocytes, and THP-1 cells (by 2.7-, 2.0-, and 1.8-fold, respectively), but did not influence coagulation in the presence of MPs from granulocytes and platelets. In accordance with these data, TF activity measured by its ability to activate factor X was found in MPs from ECs, monocytes, and THP-1 cells, but not in MPs from granulocytes and platelets. The data obtained indicate that active TF is present in MPs produced in vitro by ECs, monocytes, and THP-1 cells, but not in MPs derived from granulocytes and platelets.     

Polymorphism of uncoupling protein genes in football players: Investigation of the functional role

We studied the polymorphism of uncoupling protein genes (UCP1, UCP2, and UCP3) and the FTO gene in 28 football players (team of masters) and compared the results with the data obtained in nonathletes. All these genes encode the proteins that are involved in the regulation of body mass. However, we observed an increased percentage of the carriers of “thrifty” allele of the UCP3 gene; the alleles frequency of other studied genes showed the same tendency. This finding can be partially explained by the fact that the organism of an athlete needs to spare energy. Spearman rank correlation analysis showed that there is a significant correlation between the UCP1 and FTO genes and the elastic component of explosive strength of leg muscles. No correlations were found with the body composition and fat distribution (except for the fat distribution in the pelvic area). The UCP2 gene correlated with the parameters of respiratory functions. The UCP3 gene correlated with the rate of energy production in ramp test and the ergometric parameters of efficiency. It was also found that the high rate of uncoupling of oxidation and phosphorylation in muscles results in an increase in the total energy consumption rate, but also improves the ergometric parameters of efficiency and intensity of muscle work corresponding to the anaerobic threshold. The obtained data were analyzed in the context of the possible role of uncoupling proteins in homeostasis during intense physical activities.