100 research outputs found
Intramolecular distances between tryptophan residues and the active-site serine residue in alkaline bacterial proteinases as measured by fluorescence energy-transfer studies
Improved Mitochondrial Function with Diet-Induced Increase in Either Docosahexaenoic Acid or Arachidonic Acid in Membrane Phospholipids
Mitochondria can depolarize and trigger cell death through the opening of the mitochondrial permeability transition pore (MPTP). We recently showed that an increase in the long chain n3 polyunsaturated fatty acids (PUFA) docosahexaenoic acid (DHA; 22:6n3) and depletion of the n6 PUFA arachidonic acid (ARA; 20:4n6) in mitochondrial membranes is associated with a greater Ca2+ load required to induce MPTP opening. Here we manipulated mitochondrial phospholipid composition by supplementing the diet with DHA, ARA or combined DHA+ARA in rats for 10 weeks. There were no effects on cardiac function, or respiration of isolated mitochondria. Analysis of mitochondrial phospholipids showed DHA supplementation increased DHA and displaced ARA in mitochondrial membranes, while supplementation with ARA or DHA+ARA increased ARA and depleted linoleic acid (18:2n6). Phospholipid analysis revealed a similar pattern, particularly in cardiolipin. Tetralinoleoyl cardiolipin was depleted by 80% with ARA or DHA+ARA supplementation, with linoleic acid side chains replaced by ARA. Both the DHA and ARA groups had delayed Ca2+-induced MPTP opening, but the DHA+ARA group was similar to the control diet. In conclusion, alterations in mitochondria membrane phospholipid fatty acid composition caused by dietary DHA or ARA was associated with a greater cumulative Ca2+ load required to induced MPTP opening. Further, high levels of tetralinoleoyl cardiolipin were not essential for normal mitochondrial function if replaced with very-long chain n3 or n6 PUFAs
Systematic Review of Potential Health Risks Posed by Pharmaceutical, Occupational and Consumer Exposures to Metallic and Nanoscale Aluminum, Aluminum Oxides, Aluminum Hydroxide and Its Soluble Salts
Aluminum (Al) is a ubiquitous substance encountered both naturally (as the third most abundant element) and intentionally (used in water, foods, pharmaceuticals, and vaccines); it is also present in ambient and occupational airborne particulates. Existing data underscore the importance of Al physical and chemical forms in relation to its uptake, accumulation, and systemic bioavailability. The present review represents a systematic examination of the peer-reviewed literature on the adverse health effects of Al materials published since a previous critical evaluation compiled by Krewski et al. (2007).
Challenges encountered in carrying out the present review reflected the experimental use of different physical and chemical Al forms, different routes of administration, and different target organs in relation to the magnitude, frequency, and duration of exposure. Wide variations in diet can result in Al intakes that are often higher than the World Health Organization provisional tolerable weekly intake (PTWI), which is based on studies with Al citrate. Comparing daily dietary Al exposures on the basis of âtotal Alâassumes that gastrointestinal bioavailability for all dietary Al forms is equivalent to that for Al citrate, an approach that requires validation. Current occupational exposure limits (OELs) for identical Al substances vary as much as 15-fold.
The toxicity of different Al forms depends in large measure on their physical behavior and relative solubility in water. The toxicity of soluble Al forms depends upon the delivered dose of Al+ 3 to target tissues. Trivalent Al reacts with water to produce bidentate superoxide coordination spheres [Al(O2)(H2O4)+ 2 and Al(H2O)6 + 3] that after complexation with O2âąâ, generate Al superoxides [Al(O2âą)](H2O5)]+ 2. Semireduced AlO2âą radicals deplete mitochondrial Fe and promote generation of H2O2, O2 âą â and OHâą. Thus, it is the Al+ 3-induced formation of oxygen radicals that accounts for the oxidative damage that leads to intrinsic apoptosis. In contrast, the toxicity of the insoluble Al oxides depends primarily on their behavior as particulates.
Aluminum has been held responsible for human morbidity and mortality, but there is no consistent and convincing evidence to associate the Al found in food and drinking water at the doses and chemical forms presently consumed by people living in North America and Western Europe with increased risk for Alzheimer\u27s disease (AD). Neither is there clear evidence to show use of Al-containing underarm antiperspirants or cosmetics increases the risk of AD or breast cancer. Metallic Al, its oxides, and common Al salts have not been shown to be either genotoxic or carcinogenic. Aluminum exposures during neonatal and pediatric parenteral nutrition (PN) can impair bone mineralization and delay neurological development. Adverse effects to vaccines with Al adjuvants have occurred; however, recent controlled trials found that the immunologic response to certain vaccines with Al adjuvants was no greater, and in some cases less than, that after identical vaccination without Al adjuvants.
The scientific literature on the adverse health effects of Al is extensive. Health risk assessments for Al must take into account individual co-factors (e.g., age, renal function, diet, gastric pH). Conclusions from the current review point to the need for refinement of the PTWI, reduction of Al contamination in PN solutions, justification for routine addition of Al to vaccines, and harmonization of OELs for Al substances
Functional and Structural Properties of the 50,000 D Subunit of Octopus Vulgaris Hemocyanin
Interaction of human serum albumin with hematoporphyrin and its Zn(2)+-and Fe(3)+-derivatives.
The kinetics of the reaction of Octopus vulgaris hemocyanin with cyanide. Its significance for the structure of the 11 S subunit of molluscan hemocyanin.
Native Octopus vulgaris hemocyanin (Hc) reacts with cyanide stepwise. The first step involves the formation of a complex HcCN with O2 displacement. This complex reacts further with cyanide causing the removal of one copper ion from the active site. The same reaction sequence occurs for the extraction of the second metal ion. The formation of the HcCN complex and the removal of the first and the second copper ion can be differentiated according to the KCN concentration. The rate of metal removal is slightly affected by KCN concentration. The kinetics are dominated by site-site interactions. The kinetic curves show only slight differences when the protein is in the 11 S or 49 S aggregation states, suggesting that the site-site interactions are restricted mainly within the 11 S structure. A kinetic model describing the removal of the first copper ion is proposed assuming that the 11 S component (MW 250,000) is an annular-shaped structure made by five equivalent functional subunits (MW 50,000). An explanation for the incomplete copper removal from molluscan Hc is given. The results are compared with those previously reported for Carcinus maenas Hc
Shedding light on the mitochondrial permeability transition
The mitochondrial permeability transition is an increase of permeability of the inner mitochondrial membrane to ions and solutes with an exclusion size of about 1500 Da. It is generally accepted that the permeability transition is due to opening of a high-conductance channel, the permeability transition pore. Although the molecular nature of the permeability transition pore remains undefined, a great deal is known about its regulation and role in pathophysiology. This review specifically covers the characterization of the permeability transition pore by chemical modification of specific residues through photoirradiation of
mitochondria after treatment with porphyrins. The review also illustrates the basic principles of the photodynamic effect and the mechanisms of phototoxicity and discusses the unique properties of singlet oxygen generated by specific porphyrins in discrete mitochondrial domains. These experiments provided remarkable information on the role, interactions and topology of His and Cys residues in permeability transition pore modulation and defined an important role for the outer membrane 18 kDa translocator
protein (formerly known as the peripheral benzodiazepine receptor) in regulation of the permeability transition
The reaction between cyanide and the hemocyanin of Carcinus maenas. A kinetic study.
The kinetics of the reaction between Carcinus maenas hemocyanin and cyanide has been studied at various KCN concentrations and a different temperatures (21\ub0 and 4\ub0C) by following the decrease of the copper-peroxide absorption band, centered at 337 nm, of the copper still bound to the protein and the intrinsic fluorescence changes as functions of time. In all conditions used, the absorption band completely disappears and KCN concentration affects only the rate of the process. The reaction is kinetically homogeneous indicating no site-site interaction. The apparent rate constant increases with the square of cyanide concentration and the inverse of O2 concentration. The copper still bound decreases at a rate slower than the 337 nm absorption and the process is not kinetically homogeneous. The fluorescence of the protein increases after an induction period showing an inflection point at about 50% of the total effect. A kinetic model has been proposed on the assumption that the two metal ions are removed sequentially from the active site. The experimental data are in agreement with the theoretical equations derived from the model. The equilibrium constants for the formation of the complex between the first and the second copper ion with cyanide and the rate constants of their decomposition have been calculated. The rate-limiting process for the removal of the second copper ion is the formation of the complex with cyanide
EMISSION QUENCHING MECHANISMS IN OCTOPUS-VULGARIS HEMOCYANIN - STEADY-STATE AND TIME-RESOLVED FLUORESCENCE STUDIES
Fluorescence emission properties of various derivatives of Octopus vulgaris hemocyanin, namely, oxy, deoxy, met, half-met, half-apo, and apo derivatives, are studied by means of time-resolved and quenching techniques. Fluorescence decay can be satisfactorily fitted by two-exponential analysis in all hemocyanin derivatives. Fluorescence quenching experiments, using acrylamide, iodide, and a combination of the two, are carried out in order to correlate the observed lifetimes with different classes of fluorophores, distinguishable by their accessibility to the external quenchers. Fluorescence lifetimes of 1.2, 2.1, and 5.5 ns are attributed to buried, partially exposed, and fully exposed tryptophans, respectively, in 11s hemocyanin at pH 8.5. For 50s hemocyanin, the lifetime pattern is very similar, but a shortening of all lifetime values is observed. The fluorescence of the class of partially exposed tryptophans, situated in close proximity to the active site, is totally quenched in oxy-, met-, and half-methemocyanin. Our results rule out a Forster-type energy-transfer process from excited tryptophans to the copper-peroxide complex as a major quenching mechanism accounting for the lower quantum yield of oxyhemocyanin as compared to the deoxy and apo forms. âHeavy atomâ and âparamagnetic ionn effects, due to the bound copper, fully explain the observed finding. Thus, the application of Forsterâs theory to calculate the tryptophan active site average distance in oxyhemocyanin is not justified. Plausible values (r = 1.5 nm) are obtained by applying Forsterâs theory to the complexes of hemocyanin with carbon monoxide or 1 -anilino-8-naphthalenesulfonate
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