A protease stable in organic solvents from solvent tolerant strain of Pseudomonas aeruginosa

A solvent tolerant strain of Pseudomonas aeruginosa (PseA) was isolated from soil samples by cyclohexane enrichment in medium. The strain was able to sustain and grow in a wide range of organic solvents. The adaptation of P. aeruginosa cell towards solvents was seen at membrane level in transmission electron micrographs. It also secreted a novel protease, which exhibited remarkable solvent stability and retained most of the activity at least up to 10 days in the presence of hydrophobic organic solvents (log P > or = 2.0) at 25% (v/v) concentrations. The protease was able to withstand as high as 75% concentration of solvents at least up to 48 h. P. aeruginosa strain and its protease, both seem promising for solvent bioremediation, wastewater treatment and carrying out biotransformation in non-aqueous medium.     

Novel derivatives of ISO-1 as potent inhibitors of MIF biological function

A series of novel 1,2,4-oxadiazole, phthalimide, amide and other derivatives of ISO-1 were synthesized and probed for inhibition of macrophage migration inhibitory factor (MIF) activity. Several compounds inhibited MIF enzymatic activity at levels better than ISO-1. Of note, compounds 7, 22, 23, 24, 25 and 27 inhibited the spontaneous secretion/release/recognition of MIF from freshly isolated human peripheral blood mononuclear cells and, more importantly, inhibited the MIF-induced production of interleukin-6 (IL-6) and/or interleukin-1β (IL-1β) significantly better than ISO-1.     

Schlafen-3: A novel regulator of intestinal differentiation

Schlafen-3 (Slfn-3), a novel gene, has been shown to be a negative regulator of proliferation. The current investigation was undertaken to determine whether Slfn-3 might play a role in regulating cellular differentiation. Butyric acid, a short chain fatty acid, which induced differentiation of intestinal cells as evidenced by increased alkaline phosphatase (ALP) activity in the rat small intestinal IEC-6 cells, also produced a marked increase in Slfn-3 expression. Furthermore, overexpression of Slfn-3 caused stimulation of ALP activity in IEC-6 cells, which was exacerbated by butyrate. On the other hand, downregulation of Slfn-3 by slfn-3-si-RNA greatly attenuated the butyrate-mediated induction of differentiation of IEC-6 cells. Additionally, we observed that increased expression of Slfn-3 in colon cancer HCT-116 cells stimulated TGF-β expression and modulated expression of its downstream effectors as evidenced by increased expression of p27kip1 and downregulation of CDK-2. In addition, Slfn-3 increases E-cadherin expression but downregulates β-catenin. In conclusion, our data show that Slfn-3 plays a critical role in regulating intestinal mucosal differentiation. Furthermore our data also show that TGF-β signaling pathway plays an important role in mediating slfn-3 induced differentiation.     

Thiolato-bridged “Linear” trinuclear platinum complexes [Pt3(μ-SR)4(dppm)2]

Thiolato-bridged tri- and dinuclear platinum complexes of the types [Pt₃(μ-SR)₄(dppm)₂]²⁺ (1) and [Pt₂(μ-ER)₂(dppm)₂]²⁺ (2) (E=S or Se; R=alkyl or aryl; dppm=bis(diphenylphosphino)methane) have been prepared using the mononuclear precursors [Pt(ER)₂(dppm)]. The complexes have been characterized by NMR (¹H, ¹³C, ³¹P, ¹⁹⁵Pt), FT-IR and FAB mass spectral data. The structure of [Pt₃(μ-SC₆H₄CH₃-4)₄(dppm)₂][CF₃SO₃]₂ · 6CH₂Cl₂ (1d), has been established through X-ray crystallography, revealing a zig-zag arrangement of the three coordination spheres around the platinum atoms.     

Role of vitamin-zinc interactions on in vitro zinc uptake by human erythrocytes

In vitro zinc uptake by human erythrocytes was studied under a range of zinc concentrations representing three different plasma zinc levels i.e., zinc deficient [0.35–0.61 ppm], zinc normal [0.74–1.59 ppm], and zinc excess [1.65–2.3 ppm]. Further, interactions of physiological levels of riboflavin, flavin adenine dinucleotide (FAD), nicotinic acid, nicotinamide adenine dinucleotide (NAD), thiamine, thiamine pyrophosphate (TPP), folic acid, and ascorbic acid with zinc uptakes were studied in independent experiments. In control experiments, as compared to the normal zinc state, the rate of change of zinc uptake over change in zinc levels was 1.6 times in the excess state and 0.12 times in the deficient state, indicating three distinct patterns. Under the zinc-deficient state, thiamine significantly enhanced the zinc uptakes (p<0.05), whereas ascorbic acid and riboflavin inhibited zinc uptakes (p<0.05). The percent hemolysis of the cells was also significantly lower in the presence of thiamine (p<0.05). Under normal and excess zinc states, the vitamin-zinc interactions were not significant. The results suggest that with erythrocytes as the vehicles, thiamine might be playing an enhancer role in uptake of zinc, whereas the action of ascorbic acid might be inhibitory for zinc uptakes under deficient zinc states.     

Influence of protein flexibility on the redox potential of rubredoxin: Energy minimization studies

A theoretical investigation of the protein contribution to the redox potential of the iron–sulfur protein rubredoxin is presented. Structures of the oxidized and reduced forms of the protein were obtained by energy minimizing the oxidized crystal structure of Clostridium pasteurianum rubredoxin with appropriate charges and parameters. By including 102 crystal waters, structures close to the original crystal structure were obtained (rms difference of 1.16 Å), even with extensive minimization, thus allowing accurate calculations of comparative energies. Our calculations indicate an energy change of about –60 kcal/mol (2.58 eV) in the protein alone upon reduction. This energy change was due to both the change in charge of the redox site and the subsequent relaxation of the protein. An energy minimization procedure for the relaxation gives rms differences between the oxidized and reduced states of about 0.2 Å. The changes were small and occurred in both the backbone and sidechain mainly near the Fe–S center but contributed about – 16 kcal/mol (0.69 eV) to the total protein contribution. Although the neglect of certain effects such as electronic polarization may make the relaxation energies calculated an upper limit, the results indicate that protein relaxation contributes substantially to the redox potential. © 1993 Wiley-Liss, Inc.     

Targeted deletion of psaJ from the cyanobacterium Synechocystis sp. PCC 6803 indicates structural interactions between the PsaJ and PsaF subunits of photosystem I

Photosystem I catalyzes the light-driven oxidation of plastocyanin or cytochrome c ₆ and the reduction of ferredoxin or flavodoxin. PsaJ is a 4.4 kDa hydrophobic subunit of photosystem I from cyanobacteria and chloroplasts. To investigate the function of PsaJ, we generated a mutant strain of the cyanobacterium Synechocystis sp. PCC 6803 in which the psaJ gene is replaced by a gene for chloramphenicol resistance. Deletion of psaJ led to a reduction in the steady state RNA level from psaF which is located upstream from psaJ. Immunoquantification using an anti-PsaF antibody revealed a significant decrease in the amount of PsaF in membranes of the mutant strain. Trimeric photosystem I complexes isolated from the mutant strain using n-dodecyl -D-maltoside lacked PsaJ, contained ca. 80% less PsaF, but maintained wild-type levels of other photosystem I subunits. In contrast, the photosystem I purified using Triton X-100 contained less than 2% PsaF when compared to the wild type, showing the more extractable nature of PsaF in PsaJ-less photosystem I in the presence of Triton X-100. PsaE was more accessible to removal by NaI in a mutant strain lacking PsaF and PsaJ than in the wild type. The presence of PsaF in photosystem I from the PsaJ-less strain did not alter the increased susceptibility of PsaE to removal by NaI. These results indicate an interaction between PsaJ and PsaF in the organization of the complex.