Trimethylornithine Membrane Lipids: Discovered in Planctomycetes and Identified in Diverse Environments

Intact polar membrane lipids (IPLs) are the building blocks of all cell membranes. There is a wide range of phosphorus-free IPL structures, including amino acid containing IPLs, that can be taxonomically specific. Trimethylornithine membrane lipids (TMOs) were discovered in northern wetland Planctomycete species that were isolated and described in the last decade. The trimethylated terminal nitrogen moiety of the ornithine amino acid in the TMO structure gives the lipid a charged polar head group, similar to certain phospholipids. Since their discovery, TMOs have been identified in various other recently described northern latitude Planctomycete species, and in diverse environments including tundra soil, a boreal eutrophic lake, meso-oligotrophic lakes, and hot springs. The majority of environments or enrichment cultures in which TMOs have been observed include predominately heterotrophic microbial communities involved in the degradation of recalcitrant material and/or low oxygen methanogenic conditions at primarily northern latitudes. Other ecosystems occupied with microbial communities that possess similar metabolic pathways, such as tropical peatlands or coastal salt marshes, may include TMO producing Planctomycetes as well, further allowing these lipids to potentially be used to understand microbial community responses to environmental change in a wide range of systems. The occurrence of TMOs in hot springs indicates that these unique lipids could have broad environmental distribution with different specialized functions. Opportunities also exist to investigate the application of TMOs in microbiome studies, including forensic necrobiomes. Further environmental and microbiome lipidomics research involving TMOs will help reveal the evolution, functions, and applications of these unique membrane lipids

Identifying and Tracking Proteins Through the Marine Water Column: Insights Into the Inputs and Preservation Mechanisms of Protein in Sediments

Proteins generated during primary production represent an important fraction of marine organic nitrogen and carbon, and have the potential to provide organism-specific information in the environment. The Bering Sea is a highly productive system dominated by seasonal blooms and was used as a model system for algal proteins to be tracked through the water column and incorporated into detrital sedimentary material. Samples of suspended and sinking particles were collected at multiple depths along with surface sediments on the continental shelf and deeper basin of the Bering Sea. Modified standard proteomic preparations were used in conjunction with high pressure liquid chromatography-tandem mass spectrometry to identify the suite of proteins present and monitor changes in their distribution. In surface waters 207 proteins were identified, decreasing through the water column to 52 proteins identified in post-bloom shelf surface sediments and 24 proteins in deeper (3490 m) basin sediments. The vast majority of identified proteins in all samples were diatom in origin, reflecting their dominant contribution of biomass during the spring bloom. Identified proteins were predominantly from metabolic, binding/structural, and transport-related protein groups. Significant linear correlations were observed between the number of proteins identified and the concentration of total hydrolysable amino acids normalized to carbon and nitrogen. Organelle-bound, transmembrane, photosynthetic, and other proteins involved in light harvesting were preferentially retained during recycling. These findings suggest that organelle and membrane protection represent important mechanisms that enhance the preservation of protein during transport and incorporation into sediments

Protein Recycling in Bering Sea Algal Incubations

Protein present in phytoplankton represents a large fraction of the organic nitrogen and carbon transported from its synthesis in surface waters to marine sediments. Yet relatively little is known about the longevity of identifiable protein in situ, or the potential modifications to proteins that occur during bloom termination, protein recycling and degradation. To address this knowledge gap, diatom-dominated phytoplankton was collected during the Bering Sea spring blooms of 2009 and 2010, and incubated under darkness in separate shipboard degradation experiments spanning 11 and 53 d, respectively. In each experiment, the protein distribution was monited over time using shotgun proteomics, along with total hydrolyzable amino acids (THAAs), total protein, particulate organic carbon (POC) and nitrogen (PN), and bacterial cell abundance. Identifiable proteins, total protein and THAAs were rapidly lost during the first 5 d of enclosure in darkness in both incubations. Thereafter the loss rate was slower, and it declined further after 22 d. The initial loss of identifiable biosynthetic, glycolysis, metabolism and translation proteins after 12 h may represent shutdown of cellular activity among algal cells. Additional peptides with glycan modifications were identified in early incubation time points, suggesting that such protein modifications could be used as a marker for internal recycling processes and possibly cell death. Protein recycling was not uniform, with a subset of algal proteins including fucoxanthin chlorophyll binding proteins and RuBisCO identified after 53 d of degradation. Non-metric multidimensional scaling was used to compare the incubations with previous environmental results. The results confirmed recent observations that some fraction of algal proteins can survive water column recycling and undergo transport to marine sediments, thus contributing organic nitrogen to the benthos

Evaluation of Electrophoretic Protein Extraction and Database-Driven Protein Identification from Marine Sediments

Intact proteins comprise a major component of organic carbon and nitrogen produced globally and are likely an important fraction of organic matter in sediments and soils. Extracting the protein component from sediments and soils for mass spectral characterization and identification represents a substantial challenge given the range of products and functionalities present in the complex matrix. Multiple forms of gel electrophoresis were evaluated as a means of enhancing recovery of sedimentary protein before proteomic characterization and compared with a direct enzymatic digestion of proteins in sediments. Resulting tryptic peptides were analyzed using shotgun proteomics and tandem mass spectra were evaluated with SEQUEST. Multiple databases were then evaluated to examine the ability to confidently identify proteins from environmental samples. Following evaluation of electrophoretic extraction of proteins from sediments, the recovery of an experimentally added standard protein (BSA) from older (\u3e1 ky) sediments was optimized. Protein extraction from sediments via direct electrophoresis of a slurry mixture and the specified extraction buffer resulted in the greatest number of confident protein identifications and highest sequence coverage of the BSA standard. Searching tandem mass spectral data against larger databases with a higher diversity of proteomes did not yield a greater number of, or more confidence in, protein identifications. Regardless of the protein database used, identified peptides correlated to proteins with the same function across taxa. This suggests that while determining taxonomic-level information remains a challenge in samples with unknown mixed species, it is possible to confidently assign the function of the identified protein

Lysine and novel hydroxylysine lipids in soil bacteria: amino acid membrane lipid response to temperature and pH in <i>Pseudopedobacter saltans</i>

Microbial decomposition of organic matter is an essential process in the global carbon cycle. The soil bacteria Pseudopedobacter saltans and Flavobacterium johnsoniae are both able to degrade complex organic molecules, but it is not fully known how their membrane structures are adapted to their environmental niche. The membrane lipids of these species were extracted and analyzed using high performance liquid chromatography-electrospray ionization/ion trap/mass spectrometry (HPLC-ESI/IT/MS) and high resolution accurate mass/mass spectrometry (HRAM/MS). Abundant unknown intact polar lipids (IPLs) from P. saltans were isolated and further characterized using amino acid analysis and two dimensional nuclear magnetic resonance (NMR) spectroscopy. Ornithine IPLs (OLs) with variable (hydroxy) fatty acid composition were observed in both bacterial species. Lysine-containing IPLs (LLs) were also detected in both species and were characterized here for the first time using HPLC-MS. Novel LLs containing hydroxy fatty acids and novel hydroxylysine lipids with variable (hydroxy) fatty acid composition were identified in P. saltans. The confirmation of OL and LL formation in F. johnsoniae and P. saltans and the presence of OlsF putative homologs in P. saltans suggest the OlsF gene coding protein is possibly involved in OL and LL biosynthesis in both species, however, potential pathways of OL and LL hydroxylation in P. saltans are still undetermined. Triplicate cultures of P. saltans were grown at three temperature/pH combinations: 30°C/pH 7, 15°C/pH 7, and 15°C/pH 9. The fractional abundance of total amino acid containing IPLs containing hydroxylated fatty acids was significantly higher at higher temperature, and the fractional abundance of lysine-containing IPLs was significantly higher at lower temperature and higher pH. These results suggest that these amino acid-containing IPLs, including the novel hydroxylysine lipids, could be involved in temperature and pH stress response of soil bacteria

Agricultural data management and sharing: Best practices and case study

Agricultural data are crucial to many aspects of production, commerce, and research involved in feeding the global community. However, in most agricultural research disciplines standard best practices for data management and publication do not exist. Here we propose a set of best practices in the areas of peer review, minimal dataset development, data repositories, citizen science initiatives, and support for best data management. We illustrate some of these best practices with a case study in dairy agroecosystems research. While many common, and increasingly disparate data management and publication practices are entrenched in agricultural disciplines, opportunities are readily available for promoting and adopting best practices that better enable and enhance data-intensive agricultural research and production

Fractional quantum Hall effect in a quantum point contact at filling fraction 5/2

Recent theories suggest that the excitations of certain quantum Hall states may have exotic braiding statistics which could be used to build topological quantum gates. This has prompted an experimental push to study such states using confined geometries where the statistics can be tested. We study the transport properties of quantum point contacts (QPCs) fabricated on a GaAs/AlGaAs two dimensional electron gas that exhibits well-developed fractional quantum Hall effect, including at bulk filling fraction 5/2. We find that a plateau at effective QPC filling factor 5/2 is identifiable in point contacts with lithographic widths of 1.2 microns and 0.8 microns, but not 0.5 microns. We study the temperature and dc-current-bias dependence of the 5/2 plateau in the QPC, as well as neighboring fractional and integer plateaus in the QPC while keeping the bulk at filling factor 3. Transport near QPC filling factor 5/2 is consistent with a picture of chiral Luttinger liquid edge-states with inter-edge tunneling, suggesting that an incompressible state at 5/2 forms in this confined geometry

Measurement of the Lifetime Difference Between B_s Mass Eigenstates

We present measurements of the lifetimes and polarization amplitudes for B_s --> J/psi phi and B_d --> J/psi K*0 decays. Lifetimes of the heavy (H) and light (L) mass eigenstates in the B_s system are separately measured for the first time by determining the relative contributions of amplitudes with definite CP as a function of the decay time. Using 203 +/- 15 B_s decays, we obtain tau_L = (1.05 +{0.16}/-{0.13} +/- 0.02) ps and tau_H = (2.07 +{0.58}/-{0.46} +/- 0.03) ps. Expressed in terms of the difference DeltaGamma_s and average Gamma_s, of the decay rates of the two eigenstates, the results are DeltaGamma_s/Gamma_s = (65 +{25}/-{33} +/- 1)%, and DeltaGamma_s = (0.47 +{0.19}/-{0.24} +/- 0.01) inverse ps.Comment: 8 pages, 3 figures, 2 tables; as published in Physical Review Letters on 16 March 2005; revisions are for length and typesetting only, no changes in results or conclusion

Application of filtration blocking models to describe fouling and transmission of large plasmids DNA in sterile filtration

This is the author’s version of a work that was accepted for publication in Journal of Membrane Science. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published at: http://dx.doi.org/10.1016/j.memsci.2013.02.055Sterile filtration is considered as a final step in processing pharmaceutical grade plasmid DNA. During the development of the filtration process, fundamental understanding on the mechanism of fouling is critical to improve filtration operations. The mechanism of fouling of pQR150 (20 kb) and pGEc47 (56 kb) plasmids DNA during constant pressure filtration inside 0.22 μm PVDF membrane is experimentally investigated. The decline of filtrate flux as a function of time is analysed using the framework of classical and combined blocking models. The results for both plasmids indicate a transition between fouling mechanisms. Initially, during the early part of the filtration, the intermediate blocking model provided the best fit of the experimental results suggesting that fouling of the membrane was mainly caused by deposition of particles onto its surface. Afterwards, the result trends were best captured by the standard blocking model indicating that internal fouling of the membrane was the dominant fouling mechanism. A study of the transmission of both plasmids shows a significant reduction of plasmid transmission which coincides with the transition of the fouling mechanism from intermediate to standard blocking. The study highlights how the fouling behaviour of large plasmid DNA during sterile filtration is determined by the complex interplay between the flexibility of the molecules and the internal structure of the membrane

Refining and regaining skills in fixation/diversification stage performers: The Five-A Model

Technical change is one of many factors underpinning success in elite, fixation/diversification stage performers. Surprisingly, however, there is a dearth of research pertaining to this process or the most efficacious methods used to bring about such a change. In this paper we highlight the emergent processes, yet also the lack in mechanistic comprehension surrounding technical change, addressing issues within the motor control, sport psychology, coaching and choking literature. More importantly, we seek an understanding of how these changes can be made more secure to competitive pressure, and how this can be embedded within the process of technical change. Following this review, we propose The Five-A Model based on successful coaching techniques, psychosocial concomitants, the avoidance of choking and principles of effective behaviour change. Specific mechanisms for each stage are discussed, with a focus on the use of holistic rhythm-based cues as a possible way of internalising changes. Finally, we suggest the need for further research to examine these five stages, to aid a more comprehensive construction of the content and delivery of such a programme within the applied setting