Nonamyloid aggregates arising from mature copper/zinc superoxide dismutases resemble those observed in amyotrophic lateral sclerosis.

Protein aggregation is a hallmark of many diseases, including amyotrophic lateral sclerosis (ALS) where aggregation of copper/zinc superoxide dismutase (SOD1) is implicated in pathogenesis. We report here that fully metallated (holo) SOD1 under physiologically relevant solution conditions can undergo changes in metallation and/or dimerization over time and form aggregates that do not exhibit classical characteristics of amyloid. The relevance of the observed aggregation to disease is demonstrated by structural and tinctorial analyses, including the novel observation of binding of an anti-SOD1 antibody that specifically recognizes aggregates in ALS patients and mice models. ALS-associated SOD1 mutations can promote aggregation but are not essential. The SOD1 aggregation is characterized by a lag phase, which is diminished by self- or cross-seeding and by heterogeneous nucleation. We interpret these findings in terms of an expanded aggregation mechanism consistent with other in vitro and in vivo findings that point to multiple pathways for the formation of toxic aggregates by different forms of SOD1.

68Zn isotope exchange experiments reveal an unusual kinetic lability of the metal ions in the di-zinc form of IMP-1 metallo-beta-lactamase.

The apparently paradoxical behaviour of facile exchange (kinetic lability) of tightly bound (thermodynamic stability) zinc ions in the enzyme IMP-1 metallo-beta-lactamase with Zn-68 and cadmium ions, as indicated by in-torch vaporization inductively-coupled plasma mass spectrometry (ITV-ICP-MS) and electrospray-ionization mass spectrometry (ESI-MS), is consistent with the involvement of a third metal ion in promoting Lewis acid/base type exchange processes.

Purification and characterization of an iron-containing alcohol dehydrogenase in extremely thermophilic bacterium Thermotoga hypogea.

Thermotoga hypogea is an extremely thermophilic anaerobic bacterium capable of growing at 90 degrees C. It uses carbohydrates and peptides as carbon and energy sources to produce acetate, CO(2), H(2), L-alanine and ethanol as end products. Alcohol dehydrogenase activity was found to be present in the soluble fraction of T. hypogea. The alcohol dehydrogenase was purified to homogeneity, which appeared to be a homodimer with a subunit molecular mass of 40 +/- 1 kDa revealed by SDS-PAGE analyses. A fully active enzyme contained iron of 1.02 +/- 0.06 g-atoms/subunit. It was oxygen sensitive; however, loss of enzyme activity by exposure to oxygen could be recovered by incubation with dithiothreitol and Fe(2+). The enzyme was thermostable with a half-life of about 10 h at 70 degrees C, and its catalytic activity increased along with the rise of temperature up to 95 degrees C. Optimal pH values for production and oxidation of alcohol were 8.0 and 11.0, respectively. The enzyme had a broad specificity to use primary alcohols and aldehydes as substrates. Apparent K (m) values for ethanol and 1-butanol were much higher than that of acetaldehyde and butyraldehyde. It was concluded that the physiological role of this enzyme is likely to catalyze the reduction of aldehydes to alcohols.

Simultaneous determination of the Cd and Zn total body burden of individual, nearly microscopic, nanoliter-volume aquatic organisms ( Hyalella azteca) by rhenium-cup in-torch vaporization (ITV) sample introduction and axially viewed ICP-AES.

The Cd and Zn total body burden of individual, up to 7-day-old aquatic organisms (Hyalella azteca benthic amphipod) with an average volume of approximately 100 nL was determined simultaneously by using rhenium-cup (Re-cup) in-torch vaporization (ITV) sample introduction and an axially viewed inductively coupled plasma atomic emission spectrometry (ICP-AES) system. The direct elemental analysis capabilities of this system (i.e., no sample digestion) reduced sample preparation time, eliminated contamination concerns from the digestion reagent and, owing to its detection limits (e.g., in the low pg range for Cd and Zn), vit enabled simultaneous determinations of Cd and Zn in individual, neonate and young juvenile specimens barely visible to the unaided eye (e.g., nearly microscopic). As for calibration, liquid standards and the standard additions method were tested. Both methods gave comparable results, thus indicating that in this case liquid standards can be employed for calibration, and in the process making use of the standard additions method unnecessary. Overall, the ITV-ICP-AES approach by-passed the time-consuming acid digestions, eliminated the potential for contamination from the digestion reagents, improved considerably the speed of acquisition of analytical information and enabled simultaneous determinations of two elements using individual biological specimens.