Wargaming and NSW: Innovating for Future Conflict

Defense Analysis PosterProblem Narrative: NSW focused on landbased CT&COIN for (mostly CENTCOM) for past 15+ years; USN Fleet and NSW linkage atrophy; Re-emergence/re-focus on near-peer threats; New technology in fields of stealth, robotics, UXV, cyberNaval Special Warfare Comman

Synthesis of methylphosphonic acid by marine microbes: a source for methane in the aerobic ocean

Relative to the atmosphere, much of the aerobic ocean is supersaturated with methane; however, the source of this important greenhouse gas remains enigmatic. Catabolism of methylphosphonic acid by phosphorus-starved marine microbes, with concomitant release of methane, has been suggested to explain this phenomenon, yet methylphosphonate is not a known natural product, nor has it been detected in natural systems. Further, its synthesis from known natural products would require unknown biochemistry. Here we show that the marine archaeon Nitrosopumilus maritimus encodes a pathway for methylphosphonate biosynthesis and that it produces cell-associated methylphosphonate esters. The abundance of a key gene in this pathway in metagenomic data sets suggests that methylphosphonate biosynthesis is relatively common in marine microbes, providing a plausible explanation for the methane paradox

Radical reaction control in the AdoMet radical enzyme CDG Synthase (QueE): consolidate, destabilize, accelerate

Controlling radical intermediates and thus catalysing and directing complex radical reactions is a central feature of S-adensosylmethionine (SAM)-dependent radical enzymes. We report ab initio and DFT calculations highlighting the specific influence of ion complexation, including Mg2+, identified as a key catalytic component on radical stability and reaction control in 7-carboxy-7-deazaguanine synthase (QueE). Radical stabilisation energies (RSEs) of key intermediates and radical clock-like model systems of the enzyme-catalysed rearrangement of 6-carboxytetrahydropterin (CPH4), reveals a directing role of Mg2+ in destabilising both the substrate-derived radical and corresponding side reactions, with the effect that the experimentally-observed rearrangement becomes dominant over possible alternatives. Importantly, this is achieved with minimal disruption of the thermodynamics of the substrate itself, affording a novel mechanism for an enzyme to both maintain binding potential and accelerate the rearrangement step. Other mono and divalent ions were probed with only dicationic species achieving the necessary radical conformation to facilitate the reaction

Adenosyl Radical: Reagent and Catalyst in Enzyme Reactions

Adenosine is undoubtedly an ancient biological molecule that is a component of many enzyme cofactors: ATP, FADH, NAD(P)H, and coenzyme A, to name but a few, and, of course, of RNA. Here we present an overview of the role of adenosine in its most reactive form: as an organic radical formed either by homolytic cleavage of adenosylcobalamin (coenzyme B 12 , AdoCbl) or by single-electron reduction of S -adenosylmethionine (AdoMet) complexed to an iron–sulfur cluster. Although many of the enzymes we discuss are newly discovered, adenosine's role as a radical cofactor most likely arose very early in evolution, before the advent of photosynthesis and the production of molecular oxygen, which rapidly inactivates many radical enzymes. AdoCbl-dependent enzymes appear to be confined to a rather narrow repertoire of rearrangement reactions involving 1,2-hydrogen atom migrations; nevertheless, mechanistic insights gained from studying these enzymes have proved extremely valuable in understanding how enzymes generate and control highly reactive free radical intermediates. In contrast, there has been a recent explosion in the number of radical-AdoMet enzymes discovered that catalyze a remarkably wide range of chemically challenging reactions; here there is much still to learn about their mechanisms. Although all the radical-AdoMet enzymes so far characterized come from anaerobically growing microbes and are very oxygen sensitive, there is tantalizing evidence that some of these enzymes might be active in aerobic organisms including humans.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/69165/1/604_ftp.pd

A combined computational and experimental investigation of the [2Fe–2S] cluster in biotin synthase

Biotin synthase was the first example of what is now regarded as a distinctive enzyme class within the radical S-adenosylmethionine superfamily, the members of which use Fe/S clusters as the sulphur source in radical sulphur insertion reactions. The crystal structure showed that this enzyme contains a [2Fe–2S] cluster with a highly unusual arginine ligand, besides three normal cysteine ligands. However, the crystal structure is at such a low resolution that neither the exact coordination mode nor the role of this exceptional ligand has been elucidated yet, although it has been shown that it is not essential for enzyme activity. We have used quantum refinement of the crystal structure and combined quantum mechanical and molecular mechanical calculations to explore possible coordination modes and their influences on cluster properties. The investigations show that the protonation state of the arginine ligand has little influence on cluster geometry, so even a positively charged guanidinium moiety would be in close proximity to the iron atom. Nevertheless, the crystallised enzyme most probably contains a deprotonated (neutral) arginine coordinating via the NH group. Furthermore, the Fe···Fe distance seems to be independent of the coordination mode and is in perfect agreement with distances in other structurally characterised [2Fe–2S] clusters. The exceptionally large Fe···Fe distance found in the crystal structure could not be reproduced

Comparative Metaproteomic Analysis on Consecutively Rehmannia glutinosa-Monocultured Rhizosphere Soil

National Natural Science Foundation of China [30772729, 30671220, 31070403]; Natural Science Foundation of Fujian province, China [2008J0051]Background: The consecutive monoculture for most of medicinal plants, such as Rehmannia glutinosa, results in a significant reduction in the yield and quality. There is an urgent need to study for the sustainable development of Chinese herbaceous medicine. Methodology/Principal Findings: Comparative metaproteomics of rhizosphere soil was developed and used to analyze the underlying mechanism of the consecutive monoculture problems of R. glutinosa. The 2D-gel patterns of protein spots for the soil samples showed a strong matrix dependency. Among the spots, 103 spots with high resolution and repeatability were randomly selected and successfully identified by MALDI TOF-TOF MS for a rhizosphere soil metaproteomic profile analysis. These proteins originating from plants and microorganisms play important roles in nutrient cycles and energy flow in rhizospheric soil ecosystem. They function in protein, nucleotide and secondary metabolisms, signal transduction and resistance. Comparative metaproteomics analysis revealed 33 differentially expressed protein spots in rhizosphere soil in response to increasing years of monoculture. Among them, plant proteins related to carbon and nitrogen metabolism and stress response, were mostly up-regulated except a down-regulated protein (glutathione S-transferase) involving detoxification. The phenylalanine ammonia-lyase was believed to participate in the phenylpropanoid metabolism as shown with a considerable increase in total phenolic acid content with increasing years of monoculture. Microbial proteins related to protein metabolism and cell wall biosynthesis, were up-regulated except a down-regulated protein (geranylgeranyl pyrophosphate synthase) functioning in diterpenoid synthesis. The results suggest that the consecutive monoculture of R. glutinosa changes the soil microbial ecology due to the exudates accumulation, as a result, the nutrient cycles are affected, leading to the retardation of plant growth and development. Conclusions/Significance: Our results demonstrated the interactions among plant, soil and microflora in the proteomic level are crucial for the productivity and quality of R. glutinosa in consecutive monoculture system

Anaerobic radical enzymes for biotechnology

Enzymes that proceed through radical intermediates have a rich chemistry that includes functionalisation of otherwise unreactive carbon atoms, carbon-skeleton rearrangements, aromatic reductions, and unusual eliminations. Especially under anaerobic conditions, organisms have developed a wide range of approaches for managing these transformations that can be exploited to generate new biological routes towards both bulk and specialty chemicals. These routes are often either much more direct or allow access to molecules that are inaccessible through standard (bio)chemical approaches. This review gives an overview of some of the key enzymes in this area: benzoyl-CoA reductases (that effect the enzymatic Birch reduction), ketyl radical dehydratases, coenzyme B12-dependant enzymes, glycyl radical enzymes, and radical SAM (AdoMet radical) enzymes. These enzymes are discussed alongside biotechnological applications, highlighting the wide range of actual and potential uses. With the increased diversity in biotechnological approaches to obtaining these enzymes and information about them, even more of these amazing enzymes can be expected to find application in industrial processes