7-Carboxy-7-deazaguanine Synthase: A Radical

Radical S-adenosyl-L-methionine (SAM) enzymes are widely distributed and catalyze diverse reactions. SAM binds to the unique iron atom of a site-differentiated [4Fe-4S] cluster and is reductively cleaved to generate a 5'-deoxyadenosyl radical, which initiates turnover. 7-Carboxy-7-deazaguanine (CDG) synthase (QueE) catalyzes a key step in the biosynthesis of 7-deazapurine containing natural products. 6-Carboxypterin (6-CP), an oxidized analogue of the natural substrate 6-carboxy-5,6,7,8-tetrahydropterin (CPH4), is shown to be an alternate substrate for CDG synthase. Under reducing conditions that would promote the reductive cleavage of SAM, 6-CP is turned over to 6-deoxyadenosylpterin (6-dAP), presumably by radical addition of the 5'-deoxyadenosine followed by oxidative decarboxylation to the product. By contrast, in the absence of the strong reductant, dithionite, the carboxylate of 6-CP is esterified to generate 6-carboxypterin-5'-deoxyadenosyl ester (6-CP-dAdo ester). Structural studies with 6-CP and SAM also reveal electron density consistent with the ester product being formed in crystallo. The differential reactivity of 6-CP under reducing and nonreducing conditions highlights the ability of radical SAM enzymes to carry out both polar and radical transformations in the same active site

Radical SAM enzyme QueE defines a new minimal core fold and metal-dependent mechanism

7-carboxy-7-deazaguanine synthase (QueE) catalyzes a key S-adenosyl-L-methionine (AdoMet)- and Mg[superscript 2+]-dependent radical-mediated ring contraction step, which is common to the biosynthetic pathways of all deazapurine-containing compounds. QueE is a member of the AdoMet radical superfamily, which employs the 5′-deoxyadenosyl radical from reductive cleavage of AdoMet to initiate chemistry. To provide a mechanistic rationale for this elaborate transformation, we present the crystal structure of a QueE along with structures of pre- and post-turnover states. We find that substrate binds perpendicular to the [4Fe-4S]-bound AdoMet, exposing its C6 hydrogen atom for abstraction and generating the binding site for Mg[superscript 2+], which coordinates directly to the substrate. The Burkholderia multivorans structure reported here varies from all other previously characterized members of the AdoMet radical superfamily in that it contains a hypermodified ([β [subscript 6] over α [subscript 3]]) protein core and an expanded cluster-binding motif, CX[subscript 14]CX[subscript 2]C.United States. Dept. of Energy. Office of Biological and Environmental ResearchUnited States. Dept. of Energy. Office of Basic Energy SciencesNational Center for Research Resources (U.S.) (P41RR012408)National Institute of General Medical Sciences (U.S.) (P41GM103473)National Center for Research Resources (U.S.) (5P41RR015301-10)National Institute of General Medical Sciences (U.S.) (8 P41 GM 103403-10)United States. Dept. of Energy (Contract DE-AC02-06CH11357

Global burned area and biomass burning emissions from small fires

[1] In several biomes, including croplands, wooded savannas, and tropical forests, many small fires occur each year that are well below the detection limit of the current generation of global burned area products derived from moderate resolution surface reflectance imagery. Although these fires often generate thermal anomalies that can be detected by satellites, their contributions to burned area and carbon fluxes have not been systematically quantified across different regions and continents. Here we developed a preliminary method for combining 1-km thermal anomalies (active fires) and 500 m burned area observations from the Moderate Resolution Imaging Spectroradiometer (MODIS) to estimate the influence of these fires. In our approach, we calculated the number of active fires inside and outside of 500 m burn scars derived from reflectance data. We estimated small fire burned area by computing the difference normalized burn ratio (dNBR) for these two sets of active fires and then combining these observations with other information. In a final step, we used the Global Fire Emissions Database version 3 (GFED3) biogeochemical model to estimate the impact of these fires on biomass burning emissions. We found that the spatial distribution of active fires and 500 m burned areas were in close agreement in ecosystems that experience large fires, including savannas across southern Africa and Australia and boreal forests in North America and Eurasia. In other areas, however, we observed many active fires outside of burned area perimeters. Fire radiative power was lower for this class of active fires. Small fires substantially increased burned area in several continental-scale regions, including Equatorial Asia (157%), Central America (143%), and Southeast Asia (90%) during 2001–2010. Globally, accounting for small fires increased total burned area by approximately by 35%, from 345 Mha/yr to 464 Mha/yr. A formal quantification of uncertainties was not possible, but sensitivity analyses of key model parameters caused estimates of global burned area increases from small fires to vary between 24% and 54%. Biomass burning carbon emissions increased by 35% at a global scale when small fires were included in GFED3, from 1.9 Pg C/yr to 2.5 Pg C/yr. The contribution of tropical forest fires to year-to-year variability in carbon fluxes increased because small fires amplified emissions from Central America, South America and Southeast Asia—regions where drought stress and burned area varied considerably from year to year in response to El Nino-Southern Oscillation and other climate modes

Northern Eurasia Future Initiative (NEFI): facing the challenges and pathways of global change in the 21st century

During the past several decades, the Earth system has changed significantly, especially across Northern Eurasia. Changes in the socio-economic conditions of the larger countries in the region have also resulted in a variety of regional environmental changes that can have global consequences. The Northern Eurasia Future Initiative (NEFI) has been designed as an essential continuation of the Northern Eurasia Earth Science Partnership Initiative (NEESPI), which was launched in 2004. NEESPI sought to elucidate all aspects of ongoing environmental change, to inform societies and, thus, to better prepare societies for future developments. A key principle of NEFI is that these developments must now be secured through science-based strategies co-designed with regional decision makers to lead their societies to prosperity in the face of environmental and institutional challenges. NEESPI scientific research, data, and models have created a solid knowledge base to support the NEFI program. This paper presents the NEFI research vision consensus based on that knowledge. It provides the reader with samples of recent accomplishments in regional studies and formulates new NEFI science questions. To address these questions, nine research foci are identified and their selections are briefly justified. These foci include: warming of the Arctic; changing frequency, pattern, and intensity of extreme and inclement environmental conditions; retreat of the cryosphere; changes in terrestrial water cycles; changes in the biosphere; pressures on land-use; changes in infrastructure; societal actions in response to environmental change; and quantification of Northern Eurasia's role in the global Earth system. Powerful feedbacks between the Earth and human systems in Northern Eurasia (e.g., mega-fires, droughts, depletion of the cryosphere essential for water supply, retreat of sea ice) result from past and current human activities (e.g., large scale water withdrawals, land use and governance change) and potentially restrict or provide new opportunities for future human activities. Therefore, we propose that Integrated Assessment Models are needed as the final stage of global change assessment. The overarching goal of this NEFI modeling effort will enable evaluation of economic decisions in response to changing environmental conditions and justification of mitigation and adaptation efforts

Development and Validation of a Risk Model for Prediction of Hazardous Alcohol Consumption in General Practice Attendees: The PredictAL Study

Background: Little is known about the risk of progression to hazardous alcohol use in people currently drinking at safe limits. We aimed to develop a prediction model (predictAL) for the development of hazardous drinking in safe drinkers.Methods: A prospective cohort study of adult general practice attendees in six European countries and Chile followed up over 6 months. We recruited 10,045 attendees between April 2003 to February 2005. 6193 European and 2462 Chilean attendees recorded AUDIT scores below 8 in men and 5 in women at recruitment and were used in modelling risk. 38 risk factors were measured to construct a risk model for the development of hazardous drinking using stepwise logistic regression. The model was corrected for over fitting and tested in an external population. The main outcome was hazardous drinking defined by an AUDIT score >= 8 in men and >= 5 in women.Results: 69.0% of attendees were recruited, of whom 89.5% participated again after six months. The risk factors in the final predictAL model were sex, age, country, baseline AUDIT score, panic syndrome and lifetime alcohol problem. The predictAL model's average c-index across all six European countries was 0.839 (95% CI 0.805, 0.873). The Hedge's g effect size for the difference in log odds of predicted probability between safe drinkers in Europe who subsequently developed hazardous alcohol use and those who did not was 1.38 (95% CI 1.25, 1.51). External validation of the algorithm in Chilean safe drinkers resulted in a c-index of 0.781 (95% CI 0.717, 0.846) and Hedge's g of 0.68 (95% CI 0.57, 0.78).Conclusions: The predictAL risk model for development of hazardous consumption in safe drinkers compares favourably with risk algorithms for disorders in other medical settings and can be a useful first step in prevention of alcohol misuse

Assessment of the feasibility of a rehabilitation intervention program for breast cancer survivors with cognitive complaints

To assess the feasibility of a cognitive rehabilitation program in breast cancer survivors (BCS) with persistent post-treatment cognitive complaints. BCS with cognitive complaints, 18-months to 5-years post-treatment, were recruited for a once-weekly, five-week, group cognitive training intervention. Outcome measures included self-reported mood and cognitive function, and neurocognitive tests administered at pre-intervention, immediate-, two-month and four-month post-intervention. A sub-study in eight participants evaluated resting state quantitative electroencephalography (qEEG) changes from pre- to immediate post-intervention in relationship to post-intervention changes in cognitive complaints. Twenty-seven BCS completed the protocol and tolerated the intervention well. We observed significant reductions in total and memory-specific cognitive complaints from pre-intervention to immediate post-intervention (p = 0.031 and p = 0.009, respectively) and at four-months post-intervention (p < 0.0001 and p < 0.001, respectively). Significant improvement in neurocognitive tests were found for Symbol Digit, Stroop, and Trails A tests (df = 26, all p's <0.05). Effect sizes for changes from pre-intervention to immediate and to four-month post intervention ranged from 0.429 to 0.607, and from 0.439 to 0.741, respectively. Increase in qEEG absolute alpha power over the course of the intervention was associated with reduced complaints at immediate post-intervention (r = -0.78, p = 0.021), two-months (r range = -0.76 to -0.82, p-value range 0.004 to 0.03), and four-months (r = -0.71, p = 0.048). A five-week group cognitive training intervention is feasible and well tolerated. Cognitive complaints and neurocognitive test performances showed positive changes. qEEG may serve as a potential biomarker for improvement in self-reported complaints. A randomized clinical trial is underway to test the efficacy of the intervention

Spectroscopic, Steady-State Kinetic, and Mechanistic Characterization of the Radical SAM Enzyme QueE, Which Catalyzes a Complex Cyclization Reaction in the Biosynthesis of 7‑Deazapurines

7-Carboxy-7-deazaguanine (CDG) synthase (QueE) catalyzes the complex heterocyclic radical-mediated conversion of 6-carboxy-5,6,7,8-tetrahydropterin (CPH₄) to CDG in the third step of the biosynthetic pathway to all 7-deazapurines. Here we present a detailed characterization of QueE from <i>Bacillus subtilis</i> to delineate the mechanism of conversion of CPH₄ to CDG. QueE is a member of the radical <i>S</i>-adenosyl-l-methionine (SAM) superfamily, all of which use a bound [4Fe-4S]⁺ cluster to catalyze the reductive cleavage of the SAM cofactor to generate methionine and a 5′-deoxyadenosyl radical (5′-dAdo^•), which initiates enzymatic transformations requiring hydrogen atom abstraction. The ultraviolet–visible, electron paramagnetic resonance, and Mössbauer spectroscopic features of the homodimeric QueE point to the presence of a single [4Fe-4S] cluster per monomer. Steady-state kinetic experiments indicate a <i>K</i>_m of 20 ± 7 μM for CPH₄ and a <i>k</i>_cat of 5.4 ± 1.2 min^–1 for the overall transformation. The kinetically determined <i>K</i>_app for SAM is 45 ± 1 μM. QueE is also magnesium-dependent and exhibits a <i>K</i>_app for the divalent metal ion of 0.21 ± 0.03 mM. The SAM cofactor supports multiple turnovers, indicating that it is regenerated at the end of each catalytic cycle. The mechanism of rearrangement of QueE was probed with CPH₄ isotopologs containing deuterium at C-6 or the two prochiral positions at C-7. These studies implicate 5′-dAdo^• as the initiator of the ring contraction reaction catalyzed by QueE by abstraction of the H atom from C-6 of CPH₄