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Significance Coral reefs are highly productive ecosystems that raise a conundrum called “Darwin’s paradox”: How can high production flourish in low-nutrient conditions? We show here that in three abundant Caribbean sponges, the granules that have been commonly observed in sponge tissue for decades are polyphosphate granules. These granules can account for up to 40% of the total phosphorus (P) in sponge tissue. This finding has important implications for understanding P sequestration and recycling in the reef environment. We provide evidence that these granules are of bacterial origin and propose a P sequestration pathway by microbial symbionts and the sponge hosts. Considering the ancient origin of both partners, this process may have had an impact on the P cycle in Earth’s early history. Marine sponges are major habitat-forming organisms in coastal benthic communities and have an ancient origin in evolution history. Here, we report significant accumulation of polyphosphate (polyP) granules in three common sponge species of the Caribbean coral reef. The identity of the polyP granules was confirmed by energy-dispersive spectroscopy (EDS) and by the fluorescence properties of the granules. Microscopy images revealed that a large proportion of microbial cells associated with sponge hosts contained intracellular polyP granules. Cyanobacterial symbionts cultured from sponges were shown to accumulate polyP. We also amplified polyphosphate kinase ( ppk ) genes from sponge DNA and confirmed that the gene was expressed. Based on these findings, we propose here a potentially important phosphorus (P) sequestration pathway through symbiotic microorganisms of marine sponges. Considering the widespread sponge population and abundant microbial cells associated with them, this pathway is likely to have a significant impact on the P cycle in benthic ecosystems.

Non-haem Fe(ii)/α-ketoglutarate (αKG)-dependent enzymes harness the reducing power of αKG to catalyse oxidative reactions, usually the hydroxylation of unactivated carbons, and are involved in processes such as natural product biosynthesis, the mammalian hypoxic response, and DNA repair. These enzymes couple the decarboxylation of αKG with the formation of a high-energy ferryl-oxo intermediate that acts as a hydrogen-abstracting species. All previously structurally characterized mononuclear iron enzymes contain a 2-His, 1-carboxylate motif that coordinates the iron. The two histidines and one carboxylate, known as the 'facial triad', form one triangular side of an octahedral iron coordination geometry. A subclass of mononuclear iron enzymes has been shown to catalyse halogenation reactions, rather than the more typical hydroxylation reaction. SyrB2, a member of this subclass, is a non-haem Fe(ii)/αKG-dependent halogenase that catalyses the chlorination of threonine in syringomycin E biosynthesis. Here we report the structure of SyrB2 with both a chloride ion and αKG coordinated to the iron ion at 1.6 Å resolution. This structure reveals a previously unknown coordination of iron, in which the carboxylate ligand of the facial triad is replaced by a chloride ion.

Madagascar is well-known for the exceptional biodiversity of its macro-flora and fauna, but the biodiversity of Malagasy microbial communities remains relatively unexplored. Understanding patterns of bacterial diversity in soil and their correlations with above-ground botanical diversity could influence conservation planning as well as sampling strategies to maximize access to bacterially derived natural products. We present the first detailed description of Malagasy soil bacterial communities from a targeted 16S rRNA gene survey of greater than 290,000 sequences generated using 454 pyrosequencing. Two sampling plots in each of three forest conservation areas were established to represent different levels of disturbance resulting from human impact through agriculture and selective exploitation of trees, as well as from natural impacts of cyclones. In parallel, we performed an in-depth characterization of the total vascular plant morphospecies richness within each plot. The plots representing different levels of disturbance within each forest did not differ significantly in bacterial diversity or richness. Changes in bacterial community composition were largest between forests rather than between different levels of impact within a forest. The largest difference in bacterial community composition with disturbance was observed at the Vohibe forest conservation area, and this difference was correlated with changes in both vascular plant richness and soil pH. These results provide the first survey of Malagasy soil bacterial diversity and establish a baseline of botanical diversity within important conservation areas.

The first three-dimensional description of all the protein modules required for the activation, protection and catalytic steps of B 12 -dependent methyl transfer. Methyl transfer in a B vitamin Vitamin B 12 (cobalamin) and its derivatives are important cofactors in a broad range of biological processes, including methionine biosynthesis in humans and CO 2 fixation in acetogenic bacteria. In this manuscript, the authors structurally and biochemically characterize a large vitamin-B 12 -containing protein complex — the corrinoid iron–sulphur protein and its methyltransferase — from the model acetogen Moorella thermoacetica . The structures obtained in this study represent the first three-dimensional description of all the protein modules required for the activation, protection and catalytic steps of B 12 -dependent methyltransfer. Derivatives of vitamin B 12 are used in methyl group transfer in biological processes as diverse as methionine synthesis in humans and CO 2 fixation in acetogenic bacteria 1 , 2 , 3 . This seemingly straightforward reaction requires large, multimodular enzyme complexes that adopt multiple conformations to alternately activate, protect and perform catalysis on the reactive B 12 cofactor. Crystal structures determined thus far have provided structural information for only fragments of these complexes 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , inspiring speculation about the overall protein assembly and conformational movements inherent to activity. Here we present X-ray crystal structures of a complete 220 kDa complex that contains all enzymes responsible for B 12 -dependent methyl transfer, namely the corrinoid iron–sulphur protein and its methyltransferase from the model acetogen Moorella thermoacetica . These structures provide the first three-dimensional depiction of all protein modules required for the activation, protection and catalytic steps of B 12 -dependent methyl transfer. In addition, the structures capture B 12 at multiple locations between its ‘resting’ and catalytic positions, allowing visualization of the dramatic protein rearrangements that enable methyl transfer and identification of the trajectory for B 12 movement within the large enzyme scaffold. The structures are also presented alongside in crystallo spectroscopic data, which confirm enzymatic activity within crystals and demonstrate the largest known conformational movements of proteins in a crystalline state. Taken together, this work provides a model for the molecular juggling that accompanies turnover and helps explain why such an elaborate protein framework is required for such a simple, yet biologically essential reaction.

The sea squirt Ciona intestinalis is a well-studied model organism in developmental biology, yet little is known about its associated bacterial community. In this study, a combination of 454 pyrosequencing of 16S ribosomal RNA genes, catalyzed reporter deposition-fluorescence in situ hybridization and bacterial culture were used to characterize the bacteria living inside and on the exterior coating, or tunic, of C. intestinalis adults. The 454 sequencing data set demonstrated that the tunic bacterial community structure is different from that of the surrounding seawater. The observed tunic bacterial consortium contained a shared community of <10 abundant bacterial phylotypes across three individuals. Culture experiments yielded four bacterial strains that were also dominant groups in the 454 sequencing data set, including novel representatives of the classes Alphaproteobacteria and Flavobacteria . The relatively simple bacterial community and availability of dominant community members in culture make C. intestinalis a promising system in which to investigate functional interactions between host-associated microbiota and the development of host innate immunity.

Derivatives of vitamin [B.sub.12] are used in methyl group transfer in biological processes as diverse as methionine synthesis in humans and C[O.sub.2] fixation in acetogenic bacteria (1-3). This seemingly straightforward reaction requires large, multimodular enzyme complexes that adopt multiple conformations to alternately activate, protect and perform catalysis on the reactive [B.sub.12] cofactor. Crystal structures determined thus far have provided structural information for only fragments of these complexes (4-12), inspiring speculation about the overall protein assembly and conformational movements inherent to activity. Here we present X-ray crystal structures of a complete 220 kDa complex that contains all enzymes responsible for [B.sub.12]-dependent methyl transfer, namely the corrinoid iron-sulphur protein and its methyltransferase from the model acetogen Moorella thermoacetica. These structures provide the first three-dimensional depiction of all protein modules required for the activation, protection and catalytic steps of [B.sub.12]-dependent methyl transfer. In addition, the structures capture [B.sub.12] at multiple locations between its 'resting' and catalytic positions, allowing visualization of the dramatic protein rearrangements that enable methyl transfer and identification of the trajectory for [B.sub.12] movement within the large enzyme scaffold. The structures are also presented alongside in crystallo spectroscopic data, which confirm enzymatic activity within crystals and demonstrate the largest known conformational movements of proteins in a crystalline state. Taken together, this work provides a model for the molecular juggling that accompanies turnover and helps explain why such an elaborate protein framework is required for such a simple, yet biologically essential reaction.

Derivatives of vitamin B12 are used in methyl group transfer in biological processes as diverse as methionine synthesis in humans and CO2 fixation in acetogenic bacteria1-3. This seemingly straightforward reaction requires large, multimodular enzyme complexes that adopt multiple conformations to alternately activate, protect and perform catalysis on the reactive B12 cofactor. Crystal structures determined thus far have provided structural information for only fragments of these complexes4-12, inspiring speculation about the overall protein assembly and conformational movements inherent to activity. Here we present X-ray crystal structures of a complete 220 kDa complex that contains all enzymes responsible for B12-dependent methyl transfer, namely the corrinoid iron-sulphur protein and its methyltransferase from the model acetogen Moorella thermoacetica. These structures provide the first three-dimensional depiction of all protein modules required for the activation, protection and catalytic steps of B12-dependent methyl transfer. In addition, the structures capture B12 at multiple locations between its 'resting' and catalytic positions, allowing visualization of the dramatic protein rearrangements that enable methyl transfer and identification of the trajectory for B12 movement within the large enzyme scaffold. The structures are also presented alongside in crystallo spectroscopic data, which confirm enzymatic activity within crystals and demonstrate the largest known conformational movements of proteins in a crystalline state. Taken together, this work provides a model for the molecular juggling that accompanies turnover and helps explain why such an elaborate protein framework is required for such a simple, yet biologically essential reaction. [PUBLICATION ABSTRACT]

Non-haem Fe(ii)/alpha-ketoglutarate (alphaKG)-dependent enzymes harness the reducing power of alphaKG to catalyse oxidative reactions, usually the hydroxylation of unactivated carbons, and are involved in processes such as natural product biosynthesis, the mammalian hypoxic response, and DNA repair. These enzymes couple the decarboxylation of alphaKG with the formation of a high-energy ferryl-oxo intermediate that acts as a hydrogen-abstracting species. All previously structurally characterized mononuclear iron enzymes contain a 2- His, 1-carboxylate motif that coordinates the iron. The two histidines and one carboxylate, known as the 'facial triad', form one triangular side of an octahedral iron coordination geometry. A subclass of mononuclear iron enzymes has been shown to catalyse halogenation reactions, rather than the more typical hydroxylation reaction. SyrB2, a member of this subclass, is a non-haem Fe(ii)/alphaKG-dependent halogenase that catalyses the chlorination of threonine in syringomycin E biosynthesis. Here we report the structure of SyrB2 with both a chloride ion and alphaKG coordinated to the iron ion at 1.6 Aa resolution. This structure reveals a previously unknown coordination of iron, in which the carboxylate ligand of the facial triad is replaced by a chloride ion.

Madagascar is well-known for the exceptional biodiversity of its macro-flora and fauna, but the biodiversity of Malagasy microbial communities remains relatively unexplored. Understanding patterns of bacterial diversity in soil and their correlations with above-ground botanical diversity could influence conservation planning as well as sampling strategies to maximize access to bacterially derived natural products. We present the first detailed description of Malagasy soil bacterial communities from a targeted 16S rRNA gene survey of greater than 290,000 sequences generated using 454 pyrosequencing. Two sampling plots in each of three forest conservation areas were established to represent different levels of disturbance resulting from human impact through agriculture and selective exploitation of trees, as well as from natural impacts of cyclones. In parallel, we performed an in-depth characterization of the total vascular plant morphospecies richness within each plot. The plots representing different levels of disturbance within each forest did not differ significantly in bacterial diversity or richness. Changes in bacterial community composition were largest between forests rather than between different levels of impact within a forest. The largest difference in bacterial community composition with disturbance was observed at the Vohibe forest conservation area, and this difference was correlated with changes in both vascular plant richness and soil pH. These results provide the first survey of Malagasy soil bacterial diversity and establish a baseline of botanical diversity within important conservation areas.