An oxygen-tolerant respiratory [NiFe]-hydrogenase is proven to be a four-electron hydrogen/oxygen oxidoreductase, catalyzing the reaction 2 H 2 + O 2 = 2 H 2 O, equivalent to hydrogen combustion, over a sustained period without inactivating. At least 86% of the H 2 O produced by Escherichia coli hydrogenase-1 exposed to a mixture of 90% H 2 and 10% O 2 is accounted for by a direct four-electron pathway, whereas up to 14% arises from slower side reactions proceeding via superoxide and hydrogen peroxide. The direct pathway is assigned to O 2 reduction at the [NiFe] active site, whereas the side reactions are an unavoidable consequence of the presence of low-potential relay centers that release electrons derived from H 2 oxidation. The oxidase activity is too slow to be useful in removing O 2 from the bacterial periplasm; instead, the four-electron reduction of molecular oxygen to harmless water ensures that the active site survives to catalyze sustained hydrogen oxidation.hydrogen | mass spectrometry | Fe-S cluster H ydrogenases are enzymes that catalyze the interconversion of H 2 and H + with great efficiency. Containing Fe or Fe and Ni as active metals, they are not only important in biohydrogen production (by fermentative and photosynthetic means) but also provide inspiration for detailed understanding and development of optimal molecular electrocatalysts. The minimal active site motif, common to all hydrogenases, is a low-spin Fe atom coordinated by CO, CN − , and thiolate ligands, a combination expected to be unstable under aerobic conditions. Indeed, most hydrogenases suffer long-term or permanent inactivation when exposed to even traces of O 2 . It is therefore of special interest that certain [NiFe]-hydrogenases have evolved to sustain H 2 oxidation in the continued presence of O 2 , without inactivation: these enzymes are known as O 2 -tolerant [NiFe]-hydrogenases.Most of our current insight into the mechanism of O 2 tolerance stems from studies on respiratory membrane-bound [NiFe]-hydrogenases that couple H 2 oxidation to reduction of quinones (1-3). These enzymes are localized at the cytoplasmic membrane and project into the periplasmic space. A model proposed for the O 2 -tolerance mechanism of these [NiFe]-hydrogenases ( Fig. 1) is based on the following evidence. Oxygen reacts with O 2 -tolerant membrane-bound [NiFe]-hydrogenases to form, exclusively, an inactive state known as Ni-B or "ready," formulated as a Ni(III)-OH species, which is rapidly reactivated by one-electron transfer to rejoin the catalytic cycle of H 2 oxidation. Provided Ni-B is the sole product of O 2 attack, the presence of O 2 merely attenuates the steady-state rate of H 2 oxidation. In contrast, standard (O 2 sensitive) [NiFe]-hydrogenases react with O 2 to give a mixture of states, including ones variously known as "unready" or Ni-A, in which O 2 is either only partially reduced (possibly trapped as a peroxide) or has oxygenated atoms of the active site (3-7). The unready states are only reactivated very slowly; consequently, t...

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The extent to which climate change causes significant societal disruption remains controversial. An important example is the decline of the Akkadian Empire in northern Mesopotamia ∼4.2 ka, for which the existence of a coincident climate event is still uncertain. Here we present an Iranian stalagmite record spanning 5.2 ka to 3.7 ka, dated with 25 U/Th ages that provide an average age uncertainty of 31 y (1σ). We find two periods of increased Mg/Ca, beginning abruptly at 4.51 and 4.26 ka, and lasting 110 and 290 y, respectively. Each of these periods coincides with slower vertical stalagmite growth and a gradual increase in stable oxygen isotope ratios. The periods of high Mg/Ca are explained by periods of increased dust flux sourced from the Mesopotamia region, and the abrupt onset of this dustiness indicates threshold behavior in response to aridity. This interpretation is consistent with existing marine and terrestrial records from the broad region, which also suggest that the later, longer event beginning at 4.26 ka is of greater regional extent and/or amplitude. The chronological precision and high resolution of our record indicates that there is no significant difference, at decadal level, between the start date of the second, larger dust event and the timing of North Mesopotamia settlement abandonment, and furthermore reveals striking similarity between the total duration of the second dust event and settlement abandonment. The Iranian record demonstrates this region’s threshold behavior in dust production, and its ability to maintain this climate state for multiple centuries naturally.

Speleothem oxygen isotopes and growth rates are valuable proxies for reconstructing climate history. There is debate, however, about the conditions that allow speleothems to grow in oxygen isotope equilibrium, and about the correct equilibrium fractionation factors. We report results from a series of carbonate growth experiments in karst-analogue conditions in the laboratory. The setup closely mimics natural processes (e.g. precipitation driven by CO 2 -degassing, low ionic strength solution, thin solution film) but with a tight control on growth conditions (temperature, pCO 2 , drip rate, calcite saturation index and the composition of the initial solution). Calcite is dissolved in water in a 20,000 ppmV pCO 2 environment. This solution is dripped onto glass plates (coated with seed-carbonate) in a lower pCO 2 environment (<2500 ppmV), where degassing leads to calcite growth. Experiments were performed at 7, 15, 25 and 35°C. At each temperature, calcite was grown at three drip rates (2, 6 and 10 drips per minute) on separate plates. The mass of calcite grown in these experiments varies with temperature (T in K) and drip rate (d_r in drips min À1 ) according to the relationship daily growth mass = 1.254 + 1.478 * 10 À9 * e 0.0679T + (e 0.00248T À 2) * (À0.779d_r 2 + 10.05d_r + 11.69). This relationship indicates a substantial increase of growth mass with temperature, a smaller influence of drip rate on growth mass at low temperature and a non-linear relationship between drip rate and growth mass at higher temperatures. Low temperature, fast dripping conditions are found to be the most favourable for reducing effects associated with evaporation and rapid depletion of the dissolved inorganic carbon reservoir (rapid DIC-depletion). The impact of evaporation can be large so caves with high relative humidity are also preferable for palaeoclimate reconstruction. Even allowing for the maximum offsets that may have been induced by evaporation and rapid DIC-depletion, d 18 O measured in some of our experiments remain higher than those predicted by Kim and O'Neil (1997). Our new results are well explained by equilibrium at a significantly higher a calcite-water , with a kinetic-isotope effect that favours 16 O incorporation as growth rate increases. This scenario agrees with recent studies by Coplen (2007) and Dietzel et al. (2009). Overall, our results suggest that three separate processes cause d 18 O to deviate from true isotope equilibrium in the cave environment. Two of these drive d 18 O to higher values (evaporation and rapid DIC-depletion) while one drives d 18 O to lower values (preferential incorporation of 16 O in the solid carbonate at faster growth rates). While evaporation and DIC-depletion can be avoided in some settings, the third may be inescapable in the cave environment and means that any temperature to d 18 O relationship is an approximation. The controlled conditions of the present experiments also display limitations in the use of the Hendy test to identifying equilibrium growth.

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We have measured d 44/42 Ca of laboratory-precipitated calcite grown in an experimental setup that closely replicates stalagmite formation. Calcium solutions were dripped onto two different substrates in tightly-controlled conditions and calcite precipitated due to rapid CO 2 degassing. With seeded glass slides as the substrate, we observe a Ca isotope ratio in the calcite which is $0.5& per amu lower than that in the growth solution. This fractionation is generally almost twice that observed in previously published calcite growth experiments and indicates a large kinetic effect on Ca isotopes in the stalagmite growth environment. The precipitate forming near the spot where the drip lands shows slightly greater solution-to-precipitate fractionation than calcite further from the drip reflecting a decrease in this kinetic fractionation as precipitation continues. We interpret these results in the context of the model of Fantle and DePaolo (2007) which involves surface entrapment of light Ca isotopes to decrease calcite d 44/42 Ca, and depletion of Ca from the solution in the direct vicinity of the growing calcite to increase calcite d 44/42 Ca. In the stalagmite setting, the second of these effects is minimized so that calcite Ca isotope ratios are unusually light. This interpretation suggests that stalagmite Ca isotope ratios should decrease with the saturation state of the drip water (i.e. with the growth rate of calcite). Ca isotopes might therefore allow reconstruction of surface entrapment of trace metals and isotopes more generally and might, for instance, allow an assessment of the appropriate relationship between oxygen isotope fractionation and temperature for periods of past growth in stalagmites.