Inhibition of bovine α-chymotrypsin by cyclic trypsin inhibitor SFTI-1 isolated from sunflower seeds and its two acyclic analogues

Summary Trypsin inhibitor SFTI-1 isolated from sunflower seeds (comprising 14 amino acid residues and two cycles: head-to-tail cyclisation and disulfide bridge) is the smallest naturally occurring plant serine proteinase inhibitor. In our recent paper we have shown that the elimination head-to-tail cyclisation did not change trypsin inhibitory activity as judged by measured by association equilibrium constants K _a . The removal of disulfide bridge produced 2.4-fold lower activity. In the present paper we described chymotrypsin inhibitory activity. SFTI-1 inhibits significantly lower bovine α-chymortypsin (K _a =(5.20±1.56)×10⁶ M⁻¹). The activity of the analogue with disulfide bridge only was practically the same, whereas the K _a value determined for homodetic peptide was almost 3-fold lower. Considering the results obtained and the recent literature data we postulate the lower inhibitory activity against both enzymes of the analogue with head-to-tail cyclisation only reflect its lower proteolytic stability.     

Hydrogen cyanide and embryonal dormancy in apple seeds

Embryos of apple ( Malus domestica Borh. cv. Antonówka) were treated with 1 m M gaseous HCN for 6 h and cultured under a 12 h photoperiod. HCN pretreatment stimulated germination, increased the length of hypocotyls, shortened the main root and decreased the percentages of seedlings with asymmetrically grown as well as with asymmetrically greened cotyledons. High activity of β-cyanoalanine synthase (EC 4.4.1.9) and a sharp increase in cyanogen content during embryo culture suggested very low levels of endogenous HCN. despite the activity of HCN releasing enzymes. The obtained data allow us to postulate an important role for cyanide in the regulatory complex controlling dormancy in apple seeds. Experiments with respiratory inhibitors indicated, however, that HCN pretreatment affected neither the alternative electron transport pathway nor residual respiration.     

Characterization of the tonoplast proton pumps in Cucumis sativus L. root cells

Large-scale preparation of highly purified tonoplast from cucumber (Cucumis sativus L.) roots was obtained after centrifugation of microsome pellet (10,000 – 80,000 g) on discontinuous sucrose density gradient (20, 28, 32 and 42 %). Lack of PEP carboxylase (cytosol marker) and cytochrome c oxidase (mitochondrial marker) together with a slight activity of VO₄-ATPase (plasma membrane marker) and NADH-cytochrome c reductase (ER marker) in tonoplast preparation confirmed its high purity. Using latency of nitrate-inhibited ATPase and H⁺ pumping as criteria it was established that the majority of tonoplast vesicles were sealed and oriented right(cytoplasmic)-side-out. Strong acidification of the interior of vesicles observed at the presence of both, ATP and PP_i, confirmed that obtained tonoplast contains two classes of proton pumps: V-ATPase and H⁺PP_iase. To examine and characterise of proton-transport systems in tonoplast, the effect of various inhibitors on H⁺ pumping and hydrolytic activities of ATPase and PP_iase were measured. ATP-dependent activities (H⁺ flux and ATP hydrolysis) were specifically decreased by nitrate and bafilomycin A₁, whereas the PP_iase activities were reduced in the presence of fluoride and Na⁺ ions. Both enzymes showed a similar sensitivity to DCCD and DES. The results of experiments with KCl and NaCl suggested that the vacuolar ATPase was stimulated by Cl⁻, whereas the vacuolar Pp_iase requires K⁺ ions for its activity.     

The antioxidant properties of carnitine <Emphasis Type="Italic">in vitro</Emphasis>

Many of the effects of carnitine are ascribed to its antioxidant properties. The aim of this study was to evaluate the antioxidant properties of carnitine in vitro. Carnitine was found to decolorize ABTS^•+, and to protect fluorescein against bleaching induced by AAPH-derived peroxyl radicals and peroxynitrite, thiol groups against oxidation induced by hydrogen peroxide, peroxyl radicals, hypochlorite and peroxynitrite, and erythrocytes against hemolysis induced by peroxyl radicals and hypochlorite. These results show that carnitine has a direct antioxidant action against physiologically relevant oxidants.     

Psychological view of thyroid cancer and in hyperthyroid Graves' patients - comparative exploration

INTRODUCTION: The aim of study was the evaluation of association between psychometric factors in patients with thyroid cancer and in Graves' hyperthyroid patients. MATERIAL AND METHODS: We examined 50 patients with differentiated thyroid cancer, 42 females (84.75%), 8 males aged from 32 to 64 yr. (x +/- SD: 43 +/- 8.17 yr.) and 50 hyperthyroid patients, 45 (90%) females and 5 (10%) males corresponding aged. We used the following methods: the EAS Temperament Survey (EASD) in addition for adults, EPQ Eysenck Personality Questionnaire, Polish Abbreviated Form of the MMPI (DKO-74) and Beck's Depression Scale. We compared the scores of thyroid cancer patients and hyperthyroid patients. RESULTS: We have interpreted obtained results in both groups as similarly in: mild sense depression, high emotional control, high social dependence, ambiverssion, mild level of emotional mental balance. CONCLUSION: Results of our explorations appears great psychological similarity between differentiated thyroid patients and hyperthyroid patients.     

Polyamines in yellow lupin (<Emphasis Type="Italic">Lupinus luteus</Emphasis> L.) tolerance to soil drought

Polyamines (PAs) are related to many physiological processes, including soil drought stress. Two yellow lupin ‘Morocco 4’ (drought tolerant) and ‘Taper’ (drought sensitive) were exposed to soil drought for 2 weeks. The half of the examined plants were additionally sprayed with a solution of polyamine biosynthesis inhibitor—dl-α-difluoromethylarginine (DFMA). Yellow lupin leaves showed a 19% increase and seeds a 54% decrease in the total PA contents. The seeds contained fourfold less PAs than the leaves under drought conditions. The highest amount of spermidine and lack of agmatine were found in the leaves, while in the seeds the highest content of spermine and the presence of agmatine was confirmed. The use of DFMA under drought conditions decreased the content of spermine in ‘Morocco 4’ and ‘Taper’ (41 and 19%, respectively) and spermidine in ‘Taper’ (by 13%), as well as reduced two out of three of the yield components. More tolerant ‘Morocco 4’, after DFMA treatment was characterized by a higher spermidine and spermine content and a smaller decrease in yield components compared to the less tolerant ‘Taper’. Simultaneously subjecting plants to soil drought and DFMA treatment caused in ‘Morocco 4’ a decline in the number of pods and seeds per plant and seeds dry weight per plant (64, 50 and 54%, respectively), while in ‘Taper’ a reduction of the number of pods per plant and seeds per pod (32 and 27%, respectively) was observed. These results confirm that PAs are involved in yellow lupin tolerance and may play a protective function under soil drought conditions.     

Biological activity of pentachlorophenol on the digestive gland cells of the freshwater mussel Unio tumidus

Many chlorinated phenols and their derivatives are used extensively as insecticides, fungicides and herbicides by industrial and agricultural users throughout the world. Among these substances, pentachlorophenol (PCP) is a broad-spectrum biocide, which is still used as a wood preservative. In this paper, the digestive gland cells were used to assess the effect of PCP in the range of concentrations 3.75-75 microM (0.01-0.2 ppm) on oxidative DNA damage, fluidity changes and peroxidation activity in the plasma membrane. The toxic property of PCP on DNA strand breakage was studied using the comet assay. The results showed that pentachlorophenol in the range of 37.5-75 microM contributed to these lesions. To demonstrate the changes in the fluidity of plasma membrane we used the spectrofluorimetric method using two fluorescence probes: 1-[4-(trimethylamino)phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH) and 12-(9-anthroyloxy) stearic acid (12-AS). It was shown that PC did not influence the surface of plasma membrane but contributed to the increase in the fluidity of the internal region of the lipid bilayer in the range of concentrations 18.75-75 microM (0.05-0.2 ppm). We also examined the effect of PCP on the lipid peroxidation. To imply its peroxidation properties the spectrophotometry method was used to measure the level of malondialdehyde (MDA), one of the endpoints of the peroxidation of polyunsaturated fatty acids. The obtained results showed that PCP in the used doses did not initiate the formation of lipid peroxides. Thus, our investigation indicates that PCP can behave as a prooxidant agent but its action depends on the used doses and parameters chosen for the research.     

YbeA is the m3Psi methyltransferase RlmH that targets nucleotide 1915 in 23S rRNA

Pseudouridines in the stable RNAs of Bacteria are seldom subjected to further modification. There are 11 pseudouridine (Ψ) sites in Escherichia coli rRNA, and further modification is found only at Ψ1915 in 23S rRNA, where the N-3 position of the base becomes methylated. Here, we report the identity of the E. coli methyltransferase that specifically catalyzes methyl group addition to form m³Ψ1915. Analyses of E. coli rRNAs using MALDI mass spectrometry showed that inactivation of the ybeA gene leads to loss of methylation at nucleotide Ψ1915. Methylation is restored by complementing the knockout strain with a plasmid-encoded copy of ybeA. Homologs of the ybeA gene, and thus presumably the ensuing methylation at nucleotide m³Ψ1915, are present in most bacterial lineages but are essentially absent in the Archaea and Eukaryota. Loss of ybeA function in E. coli causes a slight slowing of the growth rate. Phylogenetically, ybeA and its homologs are grouped with other putative S-adenosylmethionine-dependent, SPOUT methyltransferase genes in the Cluster of Orthologous Genes COG1576; ybeA is the first member to be functionally characterized. The YbeA methyltransferase is active as a homodimer and docks comfortably into the ribosomal A site without encroaching into the P site. YbeA makes extensive interface contacts with both the 30S and 50S subunits to align its active site cofactor adjacent to nucleotide Ψ1915. Methylation by YbeA (redesignated RlmH for rRNA large subunit methyltransferase H) possibly functions as a stamp of approval signifying that the 50S subunit has engaged in translational initiation.     

Multispectroscopic studies of the interaction of folic acid with glycated human serum albumin

Abstract

The interaction between glycated human serum albumin (gHSA) and folic acid (FA) was investigated by various spectroscopic techniques, such as fluorescence, circular dichroism, UV–vis absorption spectroscopy and electrophoretic light scattering technique. These methods characterize the binding properties of an albumin–folic acid system. The binding constants values (K_a) at 300 and 310 K are about 10⁴ M^?1. The standard enthalpy change (ΔH) and the standard entropy change (ΔS) were calculated to be ～?20?kJ mol^?1 and ～16 J mol^?1 K^?1, respectively, which indicate characteristic electrostatic interactions between gHSA and folic acid. The CD studies showed that there are no significant conformational changes in the secondary structure of the protein. Moreover, the zeta potential measurements proved that under physiological conditions the gHSA–folic acid complex shows instability. No significant changes in the secondary structure of the protein and reversible drug binding are the desirable effect from pharmacological point of view.

Communicated by Ramaswamy H. Sarma     

Increase in visfatin after weight loss induced by gastroplastic surgery

Objective: The recently described adipokine visfatin is produced in visceral fat and has been suggested to influence insulin resistance. To investigate whether visfatin concentrations are related to changes in body weight, this adipokine was measured in insulin‐resistant severely obese patients before and after gastroplastic surgery. Research Methods and Procedures: Visfatin, interleukin‐6, high‐sensitivity C‐reactive protein, homeostasis model assessment of insulin resistance (HOMA‐IR), and other clinical parameters were assessed in 36 severely obese subjects (28 female; mean age, 43 years) with a median BMI of 44.3 kg/m² (95% confidence interval, 43.3 to 48.1 kg/m²). Results: After surgery, BMI decreased to a median of 31.9 kg/m² (30.1 to 35.1 kg/m²) (p < 0.0001). Median visfatin concentrations increased significantly after weight loss [70.9 ng/mL (61.4 to 75.6 ng/mL) vs. 86.4 ng/mL (79.4 to 89.8 ng/mL); p < 0.0005]. This increase correlated with the decrease of insulin and HOMA‐IR and was associated with a reduction in plasma interleukin‐6 and high‐sensitivity C‐reactive protein concentrations. Discussion: Massive weight loss after gastroplastic surgery is accompanied by an increase in circulating concentrations of the novel adipokine visfatin. This increase correlates with the decrease in plasma insulin concentrations and HOMA‐IR.