A U.S. Carbon Cycle Science Plan: First Meeting of the Carbon Cycle Science Working Group; Washington, D. C., 17–18 November 2008

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/95400/1/eost16754.pd

An experimental method for the in-situ observation of eutectic growth patterns in bulk samples of transparent alloys

We present an experimental method for the in-situ observation of directional-solidification fronts in bulk samples of transparent eutectic alloys. The growth front is observed obliquely in dark field through the liquid and a glass wall of the container with a long-distance microscope. We show that a focused image of the whole growth front can be obtained at a certain tilt angle of the microscope. At this tilt angle, eutectic fibers of about 3.5\mic in diameter can be clearly seen over the whole growth front in 400-\mic thick samples

A General Tool for Engineering the NAD/NADP Cofactor Preference of Oxidoreductases

The ability to control enzymatic nicotinamide cofactor utilization is critical for engineering efficient metabolic pathways. However, the complex interactions that determine cofactor-binding preference render this engineering particularly challenging. Physics-based models have been insufficiently accurate and blind directed evolution methods too inefficient to be widely adopted. Building on a comprehensive survey of previous studies and our own prior engineering successes, we present a structure-guided, semirational strategy for reversing enzymatic nicotinamide cofactor specificity. This heuristic-based approach leverages the diversity and sensitivity of catalytically productive cofactor binding geometries to limit the problem to an experimentally tractable scale. We demonstrate the efficacy of this strategy by inverting the cofactor specificity of four structurally diverse NADP-dependent enzymes: glyoxylate reductase, cinnamyl alcohol dehydrogenase, xylose reductase, and iron-containing alcohol dehydrogenase. The analytical components of this approach have been fully automated and are available in the form of an easy-to-use web tool: Cofactor Specificity Reversal–Structural Analysis and Library Design (CSR-SALAD)

Moving toward Net-Zero Emissions Requires New Alliances for Carbon Dioxide Removal

The 1.5 degrees C target will require removing at least some of the carbon dioxide (CO2) previously emitted. Knowledge on how this can be done has been increasing, though barriers remain concerning governance, policy, and acceptability. For the 26th session of the Conference of the Parties (COP26) to move beyond an academic debate on CO2 removal (CDR), a broader alliance of research and policy communities, industry, and the public is needed

A reconstruction of warm-water inflow to Upernavik Isstrøm since 1925 CE and its relation to glacier retreat

International audienceThe mass loss from the Greenland Ice Sheet has increased over the past 2 decades. Marine-terminating glaciers contribute significantly to this mass loss due to increased melting and ice discharge. Periods of rapid retreat of these tidewater glaciers have been linked to the concurrent inflow of warm Atlantic-sourced waters. However, little is known about the variability of these Atlantic-derived waters within the fjords, due to a lack of multi-annual in situ measurements. Thus, to better understand the potential role of ocean warming on glacier retreat, reconstructions that characterize the variability of Atlantic water inflow to the fjords are required. Here, we investigate foraminiferal assemblages in a sediment core from Upernavik Fjord, West Greenland, in which the major ice stream Upernavik Isstrøm terminates. We conclude that the foraminiferal assemblage is predominantly controlled by changes in bottom water composition and provide a reconstruction of Atlantic water inflow to Upernavik Fjord, spanning the period 1925–2012. This reconstruction reveals peak Atlantic water influx during the 1930s and again after 2000, a pattern that is comparable to the Atlantic Multidecadal Oscillation (AMO). The comparison of these results to historical observations of front positions of Upernavik Isstrøm reveals that inflow of warm Atlantic-derived waters likely contributed to high retreat rates in the 1930s and after 2000. However, moderate retreat rates of Upernavik Isstrøm also prevailed in the 1960s and 1970s, showing that glacier retreat continued despite a reduced Atlantic water inflow, albeit at a lower rate. Considering the link between bottom water variability and the AMO in Upernavik Fjord, and the fact that a persistent negative phase of the AMO is expected for the next decade, Atlantic water inflow into the fjord may decrease in the coming decade, potentially minimizing or stabilizing the retreat of Upernavik Isstrøm during this time interval.climate chang

A U.S. Carbon Cycle Science Plan

Report assessing the carbon cycle and providing guidance for U.S. researchers. It includes background on the history and context of the carbon cycle and previous science plan as well as chapters describing relevant fundamental science questions, science plan goals, the plan elements, interdisciplinary and international collaboration and cooperation, implementation and funding of the plan, and references with supplementary appendix information

Cofactor specificity motifs and the induced fit mechanism in class I ketol-acid reductoisomerases

Although most sequenced members of the industrially important ketol-acid reductoisomerase (KARI) family are class I enzymes, structural studies to date have focused primarily on the class II KARIs, which arose through domain duplication. In the present study, we present five new crystal structures of class I KARIs. These include the first structure of a KARI with a six-residue β2αB (cofactor specificity determining) loop and an NADPH phosphate-binding geometry distinct from that of the seven- and 12-residue loops. We also present the first structures of naturally occurring KARIs that utilize NADH as cofactor. These results show insertions in the specificity loops that confounded previous attempts to classify them according to loop length. Lastly, we explore the conformational changes that occur in class I KARIs upon binding of cofactor and metal ions. The class I KARI structures indicate that the active sites close upon binding NAD(P)H, similar to what is observed in the class II KARIs of rice and spinach and different from the opening of the active site observed in the class II KARI of Escherichia coli. This conformational change involves a decrease in the bending of the helix that runs between the domains and a rearrangement of the nicotinamide-binding site

General approach to reversing ketol-acid reductoisomerase cofactor dependence from NADPH to NADH

To date, efforts to switch the cofactor specificity of oxidoreductases from nicotinamide adenine dinucleotide phosphate (NADPH) to nicotinamide adenine dinucleotide (NADH) have been made on a case-by- case basis with varying degrees of success. Here we present a straightforward recipe for altering the cofactor specificity of a class of NADPH-dependent oxidoreductases, the ketol-acid reductoisomerases (KARIs). Combining previous results for an engineered NADH-dependent variant of Escherichia coli KARI with available KARI crystal structures and a comprehensive KARI-sequence alignment, we identified key cofactor specificity determinants and used this information to construct five KARIs with reversed cofactor preference. Additional directed evolution generated two enzymes having NADH-dependent catalytic efficiencies that are greater than the wild-type enzymes with NADPH. High-resolution structures of a wild-type/variant pair reveal the molecular basis of the cofactor switch

Artificial domain duplication replicates evolutionary history of ketol-acid reductoisomerases

The duplication of protein structural domains has been proposed as a common mechanism for the generation of new protein folds. A particularly interesting case is the class II ketol-acid reductoisomerase (KARI), which putatively arose from an ancestral class I KARI by duplication of the C-terminal domain and corresponding loss of obligate dimerization. As a result, the class II enzymes acquired a deeply embedded figure-of-eight knot. To test this evolutionary hypothesis we constructed a novel class II KARI by duplicating the C-terminal domain of a hyperthermostable class I KARI. The new protein is monomeric, as confirmed by gel filtration and x-ray crystallography, and has the deeply-knotted class II KARI fold. Surprisingly, its catalytic activity is nearly unchanged from the parent KARI. This provides strong evidence in support of domain duplication as the mechanism for the evolution of the class II KARI fold and demonstrates the ability of domain duplication to generate topological novelty in a function-neutral manner

A review of the anti-tumor potential of current therapeutics targeting the mitochondrial protease ClpP in H3K27-altered, diffuse midline glioma

Diffuse midline gliomas (DMGs) are devastating pediatric brain tumors recognized as the leading cause of cancer-related death in children. DMGs are high-grade gliomas (HGGs) diagnosed along the brain's midline. Euchromatin is the hallmark feature of DMG, caused by global hypomethylation of H3K27 either through point mutations in histone H3 genes (H3K27M), or by overexpression of the enhancer of zeste homolog inhibitory protein (EZHIP). In a clinical trial for adults with progressive HGGs, a 22-year-old patient with a thalamic H3K27-altered DMG, showed remarkable clinical and radiological responses to dordaviprone (ONC201). This response in a H3K27-altered HGG patient, coupled with the lack of response of patients harboring wildtype-H3 tumors, has increased the clinical interest in dordaviprone for the treatment of DMG. Additional reports of clinical benefit have emerged, but research defining mechanisms of action (MOA) fall behind dordaviprone's clinical use, with biomarkers of response unresolved. Here, we summarize dordaviprone's safety, interrogate its preclinical MOA- identifying the mitochondrial protease 'ClpP' as a biomarker of response, and discuss other ClpP-agonists, expanding the arsenal of potential weapons in the fight against DMG. Finally, we discuss combination strategies including ClpP-agonists, and its immunomodulatory effects suggestive of a role for the tumor microenvironment in DMG patients' response