Molecular characterization of the Hansenula polymorpha FLD1 gene encoding formaldehyde dehydrogenase

Glutathione-dependent formaldehyde dehydrogenase (FLD) is a key enzyme required for the catabolism of methanol as a carbon source and certain primary amines. such as methylamine as nitrogen sources in methylotrophic yeasts. Here we describe the molecular characterization of the FLD1 gene from the yeast Hansenula polymorpha. Unlike the recently described Pichia pastoris homologue, the H. polymorpha gene does not contain an intron. The predicted FLD1 product (Fld1p) is a protein of 380 amino acids (ca. 41 kDa) with 82% identity to P. pastoris Fld1p, 76% identity to the FLD protein sequence from n-alkane-assimilating yeast Candida maltosa and 63-64% identity to dehydrogenase class III enzymes from humans and other higher eukaryotes. The expression of FLD1 is strictly regulated and can be controlled at two expression levels by manipulation of the growth conditions. The gene is strongly induced under methylotrophic growth conditions; moderate expression is obtained under conditions in which a primary amine, e.g. methylamine, is used as nitrogen source. These properties render the FLD1 promoter of high interest for heterologous gene expression. The availability of the H. polymorpha FLD1 promoter provides an attractive alternative for expression of foreign genes besides the commonly used alcohol oxidase promoter. The sequence has been deposited in the GenBank/NCBI data library under Accession No. AF364077. Copyright (C) 2002 John Wiley Sons, Ltd.</p

A metagenomics approach for new biocatalyst discovery: application to transaminases and the synthesis of allylic amines

Transaminase enzymes have significant potential for the sustainable synthesis of amines using mild aqueous reaction conditions. Here a metagenomics mining strategy has been used for new transaminase enzyme discovery. Starting from oral cavity microbiome samples, DNA sequencing and bioinformatics analyses were performed. Subsequent in silico mining of a library of contiguous reads built from the sequencing data identified 11 putative Class III transaminases which were cloned and overexpressed. Several screening protocols were used and three enzymes selected of interest due to activities towards substrates covering a wide structural diversity. Transamination of functionalized cinnamaldehydes was then investigated for the production of valuable amine building blocks

Pre-main-sequence isochrones - III: The Cluster Collaboration isochrone server

We present an isochrone server for semi-empirical pre-main-sequence model isochrones in the following systems: Johnson–Cousins, Sloan Digital Sky Survey, Two-Micron All-Sky Survey, Isaac Newton Telescope (INT) Wide-Field Camera and INT Photometric Hα Survey (IPHAS)/UV-Excess Survey (UVEX). The server can be accessed via the Cluster Collaboration webpage http://www.astro.ex.ac.uk/people/timn/isochrones/. To achieve this, we have used the observed colours of member stars in young clusters with well-established age, distance and reddening to create fiducial loci in the colour–magnitude diagram. These empirical sequences have been used to quantify the discrepancy between the models and data arising from uncertainties in both the interior and atmospheric models, resulting in tables of semi-empirical bolometric corrections (BCs) in the various photometric systems. The model isochrones made available through the server are based on existing stellar interior models coupled with our newly derived semi-empirical BCs. As part of this analysis, we also present new cluster parameters for both the Pleiades and Praesepe, yielding ages of 135+20−11 and 665+14−7Myr as well as distances of 132 ± 2 and 184 ± 2 pc, respectively (statistical uncertainty only)

Pre-main-sequence isochrones - III. The cluster collaboration isochrone server

We present an isochrone server for semi-empirical pre-main-sequence model isochrones in the following systems: Johnson-Cousins, Sloan Digital Sky Survey, Two-Micron All-Sky Survey, Isaac Newton Telescope (INT) Wide-Field Camera and INT Photometric Ha Survey (IPHAS)/UV-Excess Survey (UVEX). The server can be accessed via the Cluster Collaboration webpage http://www.astro.ex.ac.uk/people/timn/isochrones/. To achieve this, we have used the observed colours ofmember stars in young clusters with well-established age, distance and reddening to create fiducial loci in the colour-magnitude diagram. These empirical sequences have been used to quantify the discrepancy between the models and data arising from uncertainties in both the interior and atmospheric models, resulting in tables of semi-empirical bolometric corrections (BCs) in the various photometric systems. The model isochrones made available through the server are based on existing stellar interior models coupled with our newly derived semi-empirical BCs. As part of this analysis, we also present new cluster parameters for both the Pleiades and Praesepe, yielding ages of 135+20 -11 and 665+14 -7 Myr as well as distances of 132 ± 2 and 184 ± 2 pc, respectively (statistical uncertainty only).JMR is funded by a UK Science and Technology Facilities Council (STFC) studentship. EEM acknowledges support from the National Science Foundation (NSF) Award AST-1008908. The authors would like to thank Emanuele Tognelli for the updated set of Pisa models and John Stauffer for sharing his catalogue of Kron photometric measurements of Pleiades members. The authors would also like to thank the referee for comments which have vastly improved the clarity of the manuscript. This research has made use of data obtained at the Isaac Newton Telescope, which is operated on the island of La Palma by the Isaac Newton Group (ING) in the Spanish Observatorio del Roque de los Muchachos of the Institutio de Astrofisica de Canarias. This research has made use of archival data products from the Two-Micron All-Sky Survey (2MASS), which is a joint project of the University of Massachusetts and the Infrared Processing and Analysis Center, funded by the National Aeronautics and Space Administration (NASA) and the National Science Foundation. This research has made use of public data from the SDSS. Funding for the SDSS was provided by the Alfred P. Sloan Foundation, the Participating Institutions, the National Science Foundation, the US Department of Energy, the National Aeronautics and Space Administration, the Japanese Monbukagakusho, the Max Planck Society and the Higher Education Funding Council for England. The SDSS was managed by the Astrophysical Research Consortium for the Participating Institutions

Temporal dynamics of aquatic communities and implications for pond conservation

Conservation through the protection of particular habitats is predicated on the assumption that the conservation value of those habitats is stable. We test this assumption for ponds by investigating temporal variation in macroinvertebrate and macrophyte communities over a 10-year period in northwest England. We surveyed 51 ponds in northern England in 1995/6 and again in 2006, identifying all macrophytes (167 species) and all macroinvertebrates (221 species, excluding Diptera) to species. The alpha-diversity, beta-diversity and conservation value of these ponds were compared between surveys. We find that invertebrate species richness increased from an average of 29. 5 species to 39. 8 species between surveys. Invertebrate gamma-diversity also increased between the two surveys from 181 species to 201 species. However, this increase in diversity was accompanied by a decrease in beta-diversity. Plant alpha-, beta and gamma-diversity remained approximately constant between the two periods. However, increased proportions of grass species and a complete loss of charophytes suggests that the communities are undergoing succession. Conservation value was not correlated between sampling periods in either plants or invertebrates. This was confirmed by comparing ponds that had been disturbed with those that had no history of disturbance to demonstrate that levels of correlation between surveys were approximately equal in each group of ponds. This study has three important conservation implications: (i) a pond with high diversity or high conservation value may not remain that way and so it is unwise to base pond conservation measures upon protecting currently-speciose habitats; (ii) maximising pond gamma-diversity requires a combination of late and early succession ponds, especially for invertebrates; and (iii) invertebrate and plant communities in ponds may require different management strategies if succession occurs at varying rates in the two groups

Haul-Out Behavior of Harbor Seals (Phoca vitulina) in Hood Canal, Washington

The goal of this study was to model haul-out behavior of harbor seals (Phoca vitulina) in the Hood Canal region of Washington State with respect to changes in physiological, environmental, and temporal covariates. Previous research has provided a solid understanding of seal haul-out behavior. Here, we expand on that work using a generalized linear mixed model (GLMM) with temporal autocorrelation and a large dataset. Our dataset included behavioral haul-out records from archival and VHF radio tag deployments on 25 individual seals representing 61,430 seal hours. A novel application for increased computational efficiency allowed us to examine this large dataset with a GLMM that appropriately accounts for temporal autocorellation. We found significant relationships with the covariates hour of day, day of year, minutes from high tide and year. Additionally, there was a significant effect of the interaction term hour of day : day of year. This interaction term demonstrated that seals are more likely to haul out during nighttime hours in August and September, but then switch to predominantly daylight haul-out patterns in October and November. We attribute this change in behavior to an effect of human disturbance levels. This study also examined a unique ecological event to determine the role of increased killer whale (Orcinus orca) predation on haul-out behavior. In 2003 and 2005 these harbor seals were exposed to unprecedented levels of killer whale predation and results show an overall increase in haul-out probability after exposure to killer whales. The outcome of this study will be integral to understanding any changes in population abundance as a result of increased killer whale predation

Metagenome Mining: A Sequence Directed Strategy for the Retrieval of Enzymes for Biocatalysis

Biocatalytic reactions are increasingly being used as a sustainable strategy in organic synthesis and it is recognised that there is need for new enzyme discovery. To establish the utility and versatility of a metagenomics approach, metagenomic DNA extracted from the oral cavity was sequenced and used to create an in silico contig library. This enables individual open reading frames, operons or all the enzymes of a particular family to be identified and then retrieved from the original DNA by PCR. As proof of principle a lactate dehydrogenase, a malate dehydrogenase and transketolases were identified in silico, successfully cloned and assayed. This new enzyme retrieval sequence directed method gives constructive access to metagenomic diversity and importantly improves on the low hit rate experienced when using conventional metagenomic screens

Evaluation of a fourth-generation latex agglutination test for the identification of Staphylococcus aureus

 In this study, we evaluated a fourth-generation agglutination assay (Staph Plus; DiaMondiaL[DML]) for the rapid identification of Staphylococcus aureus. First, comparison with three third-generation assays (Slidex Staph Plus, bioMérieux; Staphaurex Plus, Murex Diagnostics; Pastorex Staph-Plus, Sanofi Diagnostics Pasteur) was performed on a predefined strain collection: 265 coagulase-negative staphylococci (CNS), 266 methicillin-resistant S. aureus (MRSA) and 262 methicillin-susceptible S. aureus (MSSA) strains (“strain study”). Second, patient material-derived strains (883 CNS, 847 MSSA and 135 MRSA) were tested concurrently with both the DML and Slidex assays (“daily practice study”). In the strain study, the overall sensitivity and specificity of the DML, Slidex, Staphaurex and Pastorex assays were 99.2% and 100%, 98.1% and 100%, 95.2% and 100%, and 98.2% and 98.8%, respectively. Using the respective tests, the result was indeterminate in 0.0%, 0.6%, 0.4% and 1.5% of the strains. Overall, the sensitivity of the DML and Slidex assays were comparable in both sub-studies. However, in MRSA strains, the sensitivity of the DML assay was significantly lower than the Slidex assay. The specificity of the Slidex assay was significantly higher than the DML assay. However, the percentage of indeterminate results was much higher for the Slidex than the DML assay. In conclusion, the presumptive identification of S. aureus by the DML assay proved to be equal to third-generation latex agglutination assays

Low mass young stars in the Milky Way unveiled by DBSCAN and Gaia EDR3. Mapping the star forming regions within 1.5 Kpc

With an unprecedented astrometric and photometric data precision, Gaia EDR3 gives us, for the first time, the opportunity to systematically detect and map in the optical bands, the low mass populations of the star forming regions (SFRs) in the Milky Way. We provide a catalogue of the Gaia EDR3 data (photometry, proper motions and parallaxes) of the young stellar objects (YSOs) identified in the Galactic Plane (|b|<30 deg) within about 1.5 kpc. The catalogue of the SFRs to which they belong is also provided to study the properties of the very young clusters and put them in the context of the Galaxy structure. We applied the machine learning unsupervised clustering algorithm DBSCAN on a sample of Gaia EDR3 data photometrically selected on the region where very young stars (t<10 Myr) are expected to be found, with the aim to identify co-moving and spatially consistent stellar clusters. A subsample of 52 clusters, selected among the 7323 found with DBSCAN, has been used as template data set, to identify very young clusters from the pattern of the observed color-absolute magnitude diagrams through a pattern match process. We find 124440 candidate YSOs clustered in 354 SFRs and stellar clusters younger than 10 Myr and within about 1.5 Kpc. In addition, 65863 low mass members of 322 stellar clusters located within about 500 pc and with ages 10 Myr<t<100 Myr were also found. The selected YSOs are spatially correlated with the well known SFRs. Most of them are associated with well concentrated regions or complex structures of the Galaxy and a substantial number of them have been recognized for the first time. The massive SFRs, such as, for example, Orion, Sco-Cen and Vela, located within 600-700 pc trace a very complex three-dimensional pattern, while the farthest ones seem to follow a more regular pattern along the Galactic Plane