Search

Aims: To evaluate the ability of the haloarchaeon Haloferax volcanii to produce Ag and Au nanoparticles (NPs) and to characterize the obtained material in order to find relevant properties for future potential applications. Methods and Results: Nanoparticles were produced by incubating H. volcanii cells with the corresponding metal salt. In the presence of precursor salts, cultures evidenced a colour change associated to the formation of metallic nanostructures with plasmonic bands located in the visible range of the spectrum. X-ray fluorescence analysis confirmed the presence of Ag and Au in the NPs which were spherical, with average sizes of 25 nmol l-1 (Ag) and 10 nmol l-1 (Au), as determined by electronic microscopy. Fourier transformed infrared spectroscopy indicated that both types of NPs showed a stable protein capping. Ag NPs evidenced antibacterial activity and Au NPs improved the specificity of polymerase chain reaction reactions. Au and Ag NPs were able to reduce 4-nitrophenol when incubated with NaBH4. Conclusions: Haloferax volcanii is able to synthesize metallic NPs with interesting properties for technological applications. Significance and Impact of the Study: Our data demonstrate the ability of H. volcanii to synthesize metal NPs and constitutes a solid starting point to deepen the study and explore novel applications. [ABSTRACT FROM AUTHOR]

Aims: The effect of various nitrogen sources and nutritional starvation was examined on the production of an extracellular protease secreted by the haloalkaliphilic archaeon Natrialba magadii. Methods and Results: Cell growth and proteolytic activity were measured in cells grown with different nitrogen sources. Proteolytic activity was produced in complex and easily metabolized nitrogen sources such as yeast extract, casein and casamino acids; meanwhile, ammonium repressed enzyme production. The time course and amount of protease accumulated showed an inverse correlation with growth rate and nutrient concentration. Starvation did not induce extracellular protease production. Conclusion: The accumulation of Nab. magadii extracellular protease is stimulated by nutrient limitation and slow growth rate indicating that it is probably induced in response to a deficit in the energetic status of the cells. Nutritional starvation did not induce protease accumulation suggesting that de novo synthesis of this protease and/or factor/s necessary for its activation are required. This enzyme may be regulated by nitrogen catabolite repression and it does not require protein substrates for induction. Significance and Impact of the Study: These results contribute to the basic knowledge on protease regulation in haloalkaliphilic archaea and will help to optimize the production of this extremozyme for biotechnological applications such as protease-catalysed peptide synthesis. [ABSTRACT FROM AUTHOR]

In 2015, glyphosate was classified as "Group 2A - probably carcinogenic to humans" by the International Agency for Research on Cancer (IARC). Therefore, public concerns about the environmental and health risks of this substance have rapidly increased. Considering its toxicokinetic characteristics, urinary levels of glyphosate could be a powerful tool for human biomonitoring. Nevertheless, the physicochemical properties of this molecule and the complexity of the matrix make this purpose particularly challenging. In order to solve this problem, the presented study describes a simple LC-MS/MS method for the quantification of glyphosate in human urine after pre-column derivatization with FMOC-Cl. Method development was focused on the optimization of the derivatization reaction in human urine, adjusting critical variables such as pH of borate buffer, FMOC-Cl concentration and derivatization time. Besides, chromatographic separation and spectrometric parameters were also established. The analytical method was fully validated according international guidelines for selectivity, carry over, linearity, accuracy, precision, lower limit of quantitation, matrix effect and stability under different conditions. All performance parameters were within the acceptance criteria. In addition, the method was successfully applied to 52 urine samples obtained from exposed subjects from northern Argentina, laying the foundation for future epidemiological studies., (Copyright © 2021 Elsevier B.V. All rights reserved.)

A serine protease secreted by the haloalkaliphilic archaeon Natrialba magadii at the end of the exponential growth phase was isolated. This enzyme was purified 83 fold with a total yield of 25% by ethanol precipitation, affinity chromatography, and gel filtration. The native molecular mass of the enzyme determined by gel filtration was 45 kDa. Na. magadii extracellular protease was dependent on high salt concentrations for activity and stability, and it had an optimum temperature of 60 degrees C in the presence of 1.5M NaCl. The enzyme was stable and had a broad pH profile (6-12) with an optimum pH of 8-10 for azocasein hydrolysis. The protease was strongly inhibited by diisopropyl fluorophosphate (DFP), phenylmethyl sulfonylfluoride (PMSF), and chymostatin, indicating that it is a serine protease. It was sensitive to denaturing agents such as SDS, urea, and guanidine HCl and activated by thiol-containing reducing agents such as dithiotreitol (DTT) and 2-mercaptoethanol. This protease degraded casein and gelatin and showed substrate specificity for synthetic peptides containing Phe, Tyr, and Leu at the carboxyl terminus, showing that it has chymotrypsin-like activity. Na. magadii protease presented no cross-reactivity with polyclonal antibodies raised against the extracellular protease of Natronococcus occultus, suggesting that although these proteases share several biochemical traits, they might be antigenically unrelated.

Non-modulators are a subset of essential hypertensive individuals in whom renal hemodynamic and adrenal aldosterone responses to angiotensin II fail to modulate appropriately during high dietary salt intake. The main aim of this study was to investigate the familial aggregation of non-modulation and several erythrocyte Na+ transport systems in normotensive and hypertensive individuals as well as offspring of hypertensive parents. An additional aim was to evaluate the effect of treatment with enalapril on erythrocyte Na+ transport. We studied 15 normotensive subjects (6 males, 27+/-6 years), 14 untreated modulating essential hypertensive subjects (7 males, 38+/-7 years), 12 untreated non-modulating essential hypertensive subjects (7 males, 38+/-6 years), 14 modulating offspring of hypertensive parents (8 males, 25+/-6 years), and 14 non-modulating offspring of hypertensive parents (8 males, 26+/-4 years). Blood pressure was recorded with an oscillometric device and renal plasma flow and glomerular filtration rate by clearances of para-aminohippurate and inulin, respectively. Non-modulating subjects were identified as individuals who failed to increase effective renal plasma flow by 30% and decrease filtration fraction by at least 30% 10 days after changing from a low (20 mmol/d) to a high (250 mmol/d) sodium intake. Erythrocyte Na+ transport was characterized by measurements of the Na+-K+ pump, Na+-Li+ countertransport, Na+-K+-Cl- cotransport, passive Na+ permeability, and Na+ content. After the initial studies, hypertensive individuals were treated with enalapril (20 mg/d P.O.) for 6 months, after which erythrocyte Na+ transport measurements were again made. The main findings were that Na+-Li+ countertransport is increased in non-modulating hypertensive subjects and non-modulating offspring of hypertensive parents, that the increase in blood pressure in response to high salt intake is greater in non-modulating than modulating hypertensive subjects, and that enalapril decreases Na+-Li+ countertransport activity to normal in non-modulating hypertensive subjects. These findings provide support for a possible genetic role in the development of salt sensitivity and suggest that Na+-Li+ countertransport and non-modulation are related phenotypes.

Several abnormalities in Na metabolism have been implicated in the pathogenesis of essential hypertension. In addition, recent work by several investigators has showed that some Na transport systems in red cell membranes may be altered in those patients. In order to confirm such abnormalities we evaluated simultaneously four different and clearly defined Na transport systems in red cell membranes: the ouabain sensitive Na pump (P) and the Na-K cotransport (Co) in nystatin loaded cells, the maximal rate of Na-Li countertransport (CTT) in lithium loaded cells and the rate constant of Na passive permeability (pp) in 58 normotensive control subjects with no family history of hypertension (N), 19 normotensive subjects with family history of hypertension (NH) and 34 essential hypertensive patients (HE). The mean (mean +/- SEM microns/lc/h) value of the P and pp was found to be comparable in the three groups. Co was found significantly decreased in both HE (241 +/- 28) and in NH (227 +/- 42) when compared to the control group (290 +/- 10). Although NH also showed CTT values (377 +/- 87) higher than controls, the difference did not reach statistical significance. Our results indicate that approximately 90.9% of both HE and NH presented abnormalities in one or more of the various Na transport systems studied. Normotensive patients with a positive family history and alterations in some of the Na transport systems in red cell membranes may prove an interesting experimental model to assess the importance of such alterations for the development of hypertension.