Ligand-induced conformational changes in a thermophilic ribose-binding protein

<p>Abstract</p> <p>Background</p> <p>Members of the periplasmic binding protein (PBP) superfamily are involved in transport and signaling processes in both prokaryotes and eukaryotes. Biological responses are typically mediated by ligand-induced conformational changes in which the binding event is coupled to a hinge-bending motion that brings together two domains in a closed form. In all PBP-mediated biological processes, downstream partners recognize the closed form of the protein. This motion has also been exploited in protein engineering experiments to construct biosensors that transduce ligand binding to a variety of physical signals. Understanding the mechanistic details of PBP conformational changes, both global (hinge bending, twisting, shear movements) and local (rotamer changes, backbone motion), therefore is not only important for understanding their biological function but also for protein engineering experiments.</p> <p>Results</p> <p>Here we present biochemical characterization and crystal structure determination of the periplasmic ribose-binding protein (RBP) from the hyperthermophile <it>Thermotoga maritima </it>in its ribose-bound and unliganded state. The <it>T. maritima </it>RBP (tmRBP) has 39% sequence identity and is considerably more resistant to thermal denaturation (^<it>app</it><it>T</it>_{<it>m </it>}value is 108°C) than the mesophilic <it>Escherichia coli </it>homolog (ecRBP) (^<it>app</it><it>T</it>_{<it>m </it>}value is 56°C). Polar ligand interactions and ligand-induced global conformational changes are conserved among ecRBP and tmRBP; however local structural rearrangements involving side-chain motions in the ligand-binding site are not conserved.</p> <p>Conclusion</p> <p>Although the large-scale ligand-induced changes are mediated through similar regions, and are produced by similar backbone movements in tmRBP and ecRBP, the small-scale ligand-induced structural rearrangements differentiate the mesophile and thermophile. This suggests there are mechanistic differences in the manner by which these two proteins bind their ligands and are an example of how two structurally similar proteins utilize different mechanisms to form a ligand-bound state.</p

A major shift to the retention approach for forestry can help resolve some global forest sustainability issues

Approximately 85% of the global forest estate is neither formally protected nor in areas dedicated to intensive wood production (e.g., plantations). Given the spatial extent of unprotected forests, finding management approaches that will sustain their multiple environmental, economic, and cultural values and prevent their conversion to other uses is imperative. The major global challenge of native forest management is further demonstrated by ongoing steep declines in forest biodiversity and carbon stocks. Here, we suggest that an essential part of such management—supplementing the protection of large reserves and sensitive areas within forest landscapes (e.g., aquatic features)—is the adoption of the retention approach in forests where logging occurs. This ecological approach to harvesting provides for permanent retention of important selected structures (e.g., trees and decayed logs) to provide for continuity of ecosystem structure, function, and species composition in the postharvest forest. The retention approach supports the integration of environmental, economic, and cultural values and is broadly applicable to tropical, temperate, and boreal forests, adaptable to different management objectives, and appropriate in different societal settings. The widespread adoption of the retention approach would be one of the most significant changes in management practice since the onset of modern high-yield forestry.Fil: Lindenmayer, D.B.. The Australian National University,; AustraliaFil: Franklin, J.F.. University of Washington; Estados UnidosFil: Lõhmus, A.. University of Tartu; EstoniaFil: Baker, S.C.. University of Tasmania; AustraliaFil: Bauhus, J.. Albert Ludwigs University of Freiburg; AlemaniaFil: Beese, W.. University of Vancouver; CanadáFil: Brodie, A.. No especifíca;Fil: Kiehl, B.. Swedish University of Agricultural Sciences; SueciaFil: Kouki, J.. University of Eastern Finland; FinlandiaFil: Martínez Pastur, Guillermo José. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Austral de Investigaciones Científicas; ArgentinaFil: Messier, C.. Université du Québec a Montreal; CanadáFil: Neyland, M.. University of Tasmania; AustraliaFil: Palik, B.. No especifíca;Fil: Sverdrup Thygeson, A.. Norwegian University of Life Sciences; NoruegaFil: Volney, J.. Canadian Forest Service; CanadáFil: Wayne, A.. No especifíca;Fil: Gustafsson, L.. Swedish University of Agricultural Sciences; Sueci

Direct observation of DNA threading in flap endonuclease complexes

Maintenance of genome integrity requires that branched nucleic acid molecules are accurately processed to produce double-helical DNA. Flap endonucleases are essential enzymes that trim such branched molecules generated by Okazaki fragment synthesis during replication. Here, we report crystal structures of bacteriophage T5 flap endonuclease in complexes with intact DNA substrates, and products, at resolutions of 1.9–2.2 Å. They reveal single-stranded DNA threading through a hole in the enzyme enclosed by an inverted Vshaped helical arch straddling the active site. Residues lining the hole induce an unusual barb-like conformation in the DNA substrate juxtaposing the scissile phosphate and essential catalytic metal ions. A series of complexes and biochemical analyses show how the substrate’s single-stranded branch approaches, threads through, and finally emerges on the far side of the enzyme. Our studies suggest that substrate recognition involves an unusual “flycasting, thread, bend and barb” mechanis

Multiple novel prostate cancer susceptibility signals identified by fine-mapping of known risk loci among Europeans

Genome-wide association studies (GWAS) have identified numerous common prostate cancer (PrCa) susceptibility loci. We have fine-mapped 64 GWAS regions known at the conclusion of the iCOGS study using large-scale genotyping and imputation in 25 723 PrCa cases and 26 274 controls of European ancestry. We detected evidence for multiple independent signals at 16 regions, 12 of which contained additional newly identified significant associations. A single signal comprising a spectrum of correlated variation was observed at 39 regions; 35 of which are now described by a novel more significantly associated lead SNP, while the originally reported variant remained as the lead SNP only in 4 regions. We also confirmed two association signals in Europeans that had been previously reported only in East-Asian GWAS. Based on statistical evidence and linkage disequilibrium (LD) structure, we have curated and narrowed down the list of the most likely candidate causal variants for each region. Functional annotation using data from ENCODE filtered for PrCa cell lines and eQTL analysis demonstrated significant enrichment for overlap with bio-features within this set. By incorporating the novel risk variants identified here alongside the refined data for existing association signals, we estimate that these loci now explain ∼38.9% of the familial relative risk of PrCa, an 8.9% improvement over the previously reported GWAS tag SNPs. This suggests that a significant fraction of the heritability of PrCa may have been hidden during the discovery phase of GWAS, in particular due to the presence of multiple independent signals within the same regio

The fate of received sperm in the reproductive tract of a hermaphroditic snail and its implications for fertilisation.

Multiple mating, sperm storage and internal fertilisation enhance sperm competition. The great pond snail can use stored sperm for over three months, and frequently mates with different partners. This hermaphrodite, Lymnaea stagnalis, can also self-fertilise and often produces egg masses containing both selfed and outcrossed eggs. Hence, a sperm recipient may exert considerable control over paternity. Using microsatellite markers, we show that when allosperm are present, all genotyped eggs are cross-fertilised. We also find that sperm have the opportunity to compete, because double matings lead on average to equal paternity for each sperm donor. This indicates that received sperm are randomly mixed in storage. To gain further insight into the mechanisms underlying the process of sperm storage, digestion and utilisation, we investigated the fate of donated sperm at different times after copulation. We find that within 3 h after transfer most sperm have been transported into the sperm-digesting organ. Fluorescent labelling of sperm in histological sections further reveals that allosperm are not stored in the fertilisation pouch, but upstream in either the hermaphroditic duct, seminal vesicles, or ovotestis. Besides contributing to the understanding of the mechanisms underlying sperm competition and/or cryptic sperm choice, this study shows that mixed mating cannot be treated as a separate issue in hermaphroditic animals. © Springer Science+Business Media B.V. 2008