The hydrolytic activity of bovine adrenal medullary plasma membranes towards diadenosine polyphosphates is due to alkaline phosphodiesterase-I

AbstractA hydrolase activity directed against diadenosine 5′,5″′-P1,P4-tetraphosphate (Ap4A) has been solubilised and partially purified from the plasma membrane fraction of bovine adrenal medullary chromaffin tissue in order to determine its relationship to alkaline phosphodiesterase-I/nucleotide pyrophosphatase (PDase-I, EC 3.1.4.1). Activity with the specific dinucleoside tetraphosphatase (EC 3.6.1.17) substrate Ap4A and with the non-specific PDase-I substrate thymidine 5′-monophosphate p-nitrophenyl ester had Km and Vmax values of 2.0 μM and 600 pmol/min/mg protein and 0.2 mM and 26 nmol/min/mg protein respectively and co-chromatographed upon gel filtration and ion-exchange chromatography. Activity with the fluorescent substrates etheno-Ap4A and 4-methylumbelliferyl phenylphosphonate co-electrophoresed on native polyacrylamide gels. No activity was detected which exclusively hydrolysed Ap4A. Immunoblotting of the most purified fraction with an antibody against mouse PC-1, one of the major PDase-I family members, detected bands of 240, 120 and 62 kDa corresponding to PC-1 dimer, monomer and proteolytic fragment. Therefore, the activity previously described as bovine adrenal chromaffin cell ecto(diadenosine polyphosphate hydrolase) (ecto-ApnAase) is a PDase-I, probably bovine PC-1

Crystal structures of the GH18 domain of the bifunctional peroxiredoxin-chitinase CotE from Clostridium difficile

CotE is a coat protein that is present in the spores of Clostridium difficile, an obligate anaerobic bacterium and a pathogen that is a leading cause of antibiotic-associated diarrhoea in hospital patients. Spores serve as the agents of disease transmission, and CotE has been implicated in their attachment to the gut epithelium and subsequent colonization of the host. CotE consists of an N-terminal peroxiredoxin domain and a C-terminal chitinase domain. Here, a C-terminal fragment of CotE comprising residues 349-712 has been crystallized and its structure has been determined to reveal a core eight-stranded β-barrel fold with a neighbouring subdomain containing a five-stranded β-sheet. A prominent groove running across the top of the barrel is lined by residues that are conserved in family 18 glycosyl hydrolases and which participate in catalysis. Electron density identified in the groove defines the pentapeptide Gly-Pro-Ala-Met-Lys derived from the N-terminus of the protein following proteolytic cleavage to remove an affinity-purification tag. These observations suggest the possibility of designing peptidomimetics to block C. difficile transmission

Complementary Specificity of Unspecific Peroxygenases (UPOs) Enables Access to Diverse Products from Terpene Oxygenation

Unspecific Peroxygenases (UPOs) have emerged as attractive biocatalysts for selective oxygenations as, unlike cytochromes P450, they can be employed as easy-to-use lyophilised powders and depend only upon hydrogen peroxide as the external oxidant. The application of UPOs to a range of synthetic challenges relies on the characterisation of activity and specificity of complementary enzymes. Here we show that two UPOs, artUPO and rAaeUPO-PaDa-I-H, which are representative members of the ‘short’ Family I and ‘long’ Family II UPOs, display complementary activity in a series of scalable, preparative biotransformations of a diverse array of terpenes. The UPOs were also applied to the biotransformation of chrysanthemic acid derived fragments relevant in the agrichemical industry, culminating in the highly diastereoselective and enantioselective oxidation of a racemic synthetic pyrethroid derivative via kinetic resolution

The Right Light – De Novo Design of a Robust Modular Photochemical Reactor for Optimum Batch and Flow Chemistry

Having identified inconsistencies when repeating literature examples of photochemical transformations and difficulties recreating experimental setups, we devised several criteria that an ideal labscale reactor should achieve. Herein, we introduce a versatile photoreactor for high throughput screening, preparative scale batch reactions and continuous processing, all with a single light source. The reactor utilizes interchangeable arrays of pseudo-monochromatic high-power LEDs in a range of synthetically useful wavelengths, combined with excellent temperature control. Moreover, light intensity can be modulated in an accurate and straightforward manner. This system has subsequently been tested on a range of literature methodologies

A community-maintained standard library of population genetic models

The explosion in population genomic data demands ever more complex modes of analysis, and increasingly, these analyses depend on sophisticated simulations. Recent advances in population genetic simulation have made it possible to simulate large and complex models, but specifying such models for a particular simulation engine remains a difficult and error-prone task. Computational genetics researchers currently re-implement simulation models independently, leading to inconsistency and duplication of effort. This situation presents a major barrier to empirical researchers seeking to use simulations for power analyses of upcoming studies or sanity checks on existing genomic data. Population genetics, as a field, also lacks standard benchmarks by which new tools for inference might be measured. Here, we describe a new resource, stdpopsim, that attempts to rectify this situation. Stdpopsim is a community-driven open source project, which provides easy access to a growing catalog of published simulation models from a range of organisms and supports multiple simulation engine backends. This resource is available as a well-documented python library with a simple command-line interface. We share some examples demonstrating how stdpopsim can be used to systematically compare demographic inference methods, and we encourage a broader community of developers to contribute to this growing resource.Open access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

The khmer software package: enabling efficient nucleotide sequence analysis

The khmer package is a freely available software library for working efficiently with fixed length DNA words, or k-mers. khmer provides implementations of a probabilistic k-mer counting data structure, a compressible De Bruijn graph representation, De Bruijn graph partitioning, and digital normalization. khmer is implemented in C++ and Python, and is freely available under the BSD license at https://github.com/dib-lab/khmer/

The khmer software package: enabling efficient nucleotide sequence analysis [version 1; referees: 2 approved, 1 approved with reservations]

The khmer package is a freely available software library for working efficiently with fixed length DNA words, or k-mers. khmer provides implementations of a probabilistic k-mer counting data structure, a compressible De Bruijn graph representation, De Bruijn graph partitioning, and digital normalization. khmer is implemented in C++ and Python, and is freely available under the BSD license at https://github.com/dib-lab/khmer/