Use of UV endonuclease from Micrococcus luteus to monitor the progress of DNA repair in UV-irradiated human cells

A sensitive enzymatic assay has been developed to follow the progress of NDA repair in human cells exposed to UV radiation. The assay employs an endonuclease selectively active at sites containing pyrimidine dimers in UV-damaged DNA. Primary fibroblasts are exposed to 254 nm radiation and incubated for specified times, their radioactivity labelled DNA is isolated and treated with a UV endonuclease extensively purified from Micrococcus luteus. Endonuclease-susceptible site remaining in the DNA are subsequently observed as single-strand scissions by sedimentation in alkaline sucrose gradients. In comparison to the situation with excision-proficient normal cells, those derived from patients suffering from either the classical or the De Sanctis-Cacchione clinical form of Xeroderma pigmentosum (XP) exhibit a marked diminution in the rate of disappearance of nuclease-susceptible lesions with time of post-UV incubation.     

A manganese(II) dichloro-bridged one-dimensional polymer: Structural, fluorescence and low-temperature magnetic study

One-dimensional organic/inorganic composite coordination polymer has been synthesised by the reaction of manganese(II) chloride with the chelating bidentate ligand, 1,10-phenanthroline (1,10-phen). X-ray single crystal analysis shows a doubly chloride bridged 1-D polymer, [Mn(μ-Cl)₂(phen)]_n (1), where manganese(II) ions possess octahedral environment. The complex is characterised by elemental analysis, different spectroscopic, electrochemical and low temperature magnetic susceptibility measurements. 1 exhibits strong fluorescence emission band at 410 nm and can serve as potential photoactive material as indicated from the characteristic fluorescence properties. Magnetic susceptibility measurements reveal a weak ferromagnetic interaction between the two high-spin Mn(II) ions of J = 0.017 cm⁻¹.     

Enhancement of exciton coupled circular dichroism with sterically encumbered bis-porphyrin tweezers

Porphyrin tweezers have been successfully used as hosts for the absolute stereochemical determination of a variety of chiral compounds. A set of new porphyrin tweezers with substituted aryl groups on the meso position of the porphyrin rings have been synthesized. The modified tweezers are used as hosts for the stereochemical determination of chiral diamines and carrier-derivatized alpha-chiral carboxylic acids in order to monitor the influence of the various substitutions of the aryl group on the amplitude and sign of the ECCD couplet. t-Butyl substitution at the meta positions of the porphyrin's meso phenyl substituents leads to enhanced ECCD amplitudes.     

Synthesis, spectroscopic and structural characterization of the high-spin Fe(II) cyanato-N and thiocyanato-N “picket fence” porphyrin complexes

Two new iron(II) five-coordinated porphyrin complexes [Na(2,2,2-crypt)] [Fe^II(TpivPP)(NCO)] (1) (TpivPP = α,α,α,α-tetrakis(o-pivalamidophenyl) porphyrin known as picket fence porphyrin and 2,2,2-crypt is the cryptand-222) and [K(2,2,2-crypt)][Fe^II(TpivPP)(NCS)] (2) have been prepared and characterized. The UV-Vis and IR spectroscopic data are consistent with a cyanato-N and thiocyanato-N ferrous porphyrinates. The Mössbauer data and the X-ray structural analysis indicate that the Fe(II) cation in 1 and 2 is high-spin (S = 2) and has the (d_xy)²(d_xz)¹(d_yz)¹(d_z²)¹(d_x²-y²)¹ ground state electronic configuration.For complex 1, the average equatorial iron-pyrrole N bond length (Fe-N_p = 2.120(2) Å), the distance between the iron and the 24-atom mean plane of the porphyrin ring (Fe-P_C = 0.6805(7) Å) and the distance between the iron and the plane made by the four pyrrole nitrogens (Fe-P_N = 0.5923(12) Å) are longer than those of complex 2 and similar five-coordinated Fe(II) high-spin porphyrinates. This is probably due to the significant electronic repulsion of the d_x²-y² and d_xy orbitals by the negative charge of the pyrrole N atoms in case of 1.     

Pot size affects expression of Mn efficiency in barley

Mn efficiency is defined here as an ability of a genotype to grow and yield well in a soil which is limiting in available Mn for a standard genotype (Graham, 1984). Screening for Mn efficiency in soil-based pot testing had been producing inconsistent results, and thus improvement of pot screening became an objective. One possible factor, pot size was examined as the cause, using two sizes of pot. In large pots, the expectation of higher dry matter and shoot Mn concentration in a Mn-efficient genotype compared to a Mn-inefficient genotype was realised over a wide range of Mn supply, whereas in small pots, the genotypic differences were expressed at only one, low rate of Mn supply (10 mg kg soil^-1). Plants in the small pots strongly responded to root restriction by decreasing yields and increasing root/shoot ratios and Mn concentrations of shoots. The critical value of Mn concentration for shoot growth was not affected by the small pots, but the Mn mobilization by plants might be affected in the small pots. The practical outcome of these results is that using an adequate size of pot and measuring the Mn concentration of shoots, soil-based pot screening for Mn efficiency can be improved.     

Metals (Fe, Mn, Zn) in the root plaque of submerged aquatic plants collected in situ: Relations with metal concentrations in the adjacent sediments and in the root tissue

We have investigated the extent of iron oxyhydroxide deposition on the roots of two common freshwater species, Vallisneria americana Michx. and Heteranthera dubia (Jacq.) MacM., collected from different sites in the St. Lawrence River, Québec, Canada, and have related metal concentrations in the root plaques both to the geochemical conditions prevailing in the host sediments (pH; metal partitioning) and to the metal concentrations within the plant root tissue. Possible effects of root plaque on sediment geochemistry are also discussed.At those sites where the two submerged plants co-existed, the amounts of Fe deposited on their respective root surfaces were positively correlated, indicating that sediment geochemistry (pH; concentration of labile metal) exerted a more important influence on plaque formation than did inter-species differences (root physiology, morphology). Iron and Mn concentrations in the root plaque were positively correlated with each other, and with the readily extractable fractions (F1, 172) of these metals in the adjacent sediments. In contrast, Zn concentrations in the root plaque of V. americana were not related to Zn concentrations in the sediments — the dominant geochemical process at the root surface is Fe deposition, such that the quantities of Zn deposited on the roots are determined not by Zn geochemistry per se but rather by the amount of Fe deposition. Indeed the Zn/Fe ratios in the root plaque were related to the Zn/Fe ratios in the surrounding sediments (NH₂OHHCl extract).On a concentration basis (g/g), more Fe, Mn and Zn was found outside the root, in the iron plaque, than inside the root tissues. For all 3 metals, significant relationships were observed between the metal concentrations in the plaque and those inside the roots. For Zn, however, the best statistical relationship was not with [Zn]_plaque, but rather with the [Zn]/[Fe] ratio in the plaque. It is hypothesized that the Zn/Fe ratio in the root plaque reflects the free Zn²⁺ concentration adjacent to the root surface, and that this in turn affects Zn uptake by the plant root. For a given value of Zn in the sediments or in the root plaque, the Zn content of the root is inversely related to the concentration of Fe oxyhydroxides, implying that Fe plays a protective role in regulating Zn bioavailability.     

Physiological and proteomic characterization of manganese sensitivity and tolerance in rice (Oryza sativa) in comparison with barley (Hordeum vulgare)

Background and Aims

Research on manganese (Mn) toxicity and tolerance indicates that Mn toxicity develops apoplastically through increased peroxidase activities mediated by phenolics and Mn, and Mn tolerance could be conferred by sequestration of Mn in inert cell compartments. This comparative study focuses on Mn-sensitive barley (Hordeum vulgare) and Mn-tolerant rice (Oryza sativa) as model organisms to unravel the mechanisms of Mn toxicity and/or tolerance in monocots.

Methods

Bulk leaf Mn concentrations as well as peroxidase activities and protein concentrations were analysed in apoplastic washing fluid (AWF) in both species. In rice, Mn distribution between leaf compartments and the leaf proteome using 2D isoelectic focusing IEF/SDS–PAGE and 2D Blue native BN/SDS–PAGE was studied.

Key Results

The Mn sensitivity of barley was confirmed since the formation of brown spots on older leaves was induced by low bulk leaf and AWF Mn concentrations and exhibited strongly enhanced H₂O₂-producing and consuming peroxidase activities. In contrast, by a factor of 50, higher Mn concentrations did not produce Mn toxicity symptoms on older leaves in rice. Peroxidase activities, lower by a factor of about 100 in the rice leaf AWF compared with barley, support the view of a central role for these peroxidases in the apoplastic expression of Mn toxicity. The high Mn tolerance of old rice leaves could be related to a high Mn binding capacity of the cell walls. Proteomic studies suggest that the lower Mn tolerance of young rice leaves could be related to Mn excess-induced displacement of Mg and Fe from essential metabolic functions.

Conclusions

The results provide evidence that Mn toxicity in barley involves apoplastic lesions mediated by peroxidases. The high Mn tolerance of old leaves of rice involves a high Mn binding capacity of the cell walls, whereas Mn toxicity in less Mn-tolerant young leaves is related to Mn-induced Mg and Fe deficiencies.     

Removal of Mn(II) ions from aqueous neutral media by manganese-oxidizing fungus in the presence of carbon fiber

A manganese-oxidizing fungus was isolated from a hot spring in Japan. The fungus was increasingly effective at oxidizing Mn(II) ions as the concentration of organic carbon sources in the growth medium was decreased. The fungus oxidized 50 ppm of Mn(II) ions within 160 h in a pH 7.3 medium at 25 degrees C. The presence of carbon fiber shortened the time to 80 h, and promoted steady oxidation. The oxidation products were identified by XPS and XRD to be poorly crystallized and amorphous MnO(2), both with and without the fiber. These results suggest that the fiber participates in kinetically limited oxidation. The fungus was entangled with and clung to the fibers, and the oxidized Mn species accumulated on the fungus. Similarly shaped polyethylene telephthalate fiber did not enhance the oxidation, nor was adhesion of the fungus observed. Although the mechanism is still unknown, the present work shows that removal of Mn from solution through the precipitation and accumulation of Mn-oxides on the fungus in the presence of carbon fiber is a promising improvement for water treatment.