Structure‐Based Mechanism of Oleate Hydratase from Elizabethkingia meningoseptica

Hydratases provide access to secondary and tertiary alcohols by regio‐ and/or stereospecifically adding water to carbon‐carbon double bonds. Thereby, hydroxy groups are introduced without the need for costly cofactor recycling, and that makes this approach highly interesting on an industrial scale. Here we present the first crystal structure of a recombinant oleate hydratase originating from Elizabethkingia meningoseptica in the presence of flavin adenine dinucleotide (FAD). A structure‐based mutagenesis study targeting active site residues identified E122 and Y241 as crucial for the activation of a water molecule and for protonation of the double bond, respectively. Moreover, we also observed that two‐electron reduction of FAD results in a sevenfold increase in the substrate hydration rate. We propose the first reaction mechanism for this enzyme class that explains the requirement for the flavin cofactor and the involvement of conserved amino acid residues in this regio‐ and stereoselective hydration.     

Synthesis,morphology and properties of segmented poly(ether ester amide)s comprising uniform glycine or β-alanine extended bisoxalamide hard segments

Segmented poly(ether ester amide)s comprising glycine or β-alanine extended bisoxalamide hard segments are highly phase separated thermoplastic elastomers with a broad temperature independent rubber plateau. These materials with molecular weights, M_n, exceeding 30 × 10³ g mol^?1 are conveniently prepared by polycondensation of preformed bisester–bisoxalamides and commercially available PTHF diols. FT-IR revealed strongly hydrogen bonded and highly ordered bisoxalamide hard segments with degrees of ordering between 73 and 99%. The morphology consists of fiber-like nano-crystals randomly dispersed in the soft polymer matrix. The micro-structural parameters of the copolymers were addressed by simultaneous small- and wide-angle X-ray scattering. It is shown that the crystals have strictly identical thickness, which is close to the contour length of the hard segment. The long dimension of the crystals is identified with the direction of the hydrogen bonds. The melting transitions of the hard segments are sharp, with temperatures up to 170 °C. The studied polymers have an elastic modulus in the range of 139–170 MPa, a stress at break in the range of 19–31 MPa combined with strains at break of higher than 800%. The segmented copolymer comprising the β-alanine based bisoxalamide hard segment with a spacer of 6 methylene groups has a melting transition of 141 °C which is higher than the melting transition of its glycine analogue of 119 °C. Likewise, the fracture stress increased from 22 to 31 MPa when the glycine ester group in the hard segment was replaced with β-alanine. The improved thermal and mechanical properties of the latter polymers is related to the crystal packing of the β-alanine based hard segments in the copolymer compared to the packing of the hard segments comprising glycine ester groups.     

Chemical-Shift Perturbations Reflect Bile Acid Binding to Norovirus Coat Protein: Recognition Comes in Different Flavors

Bile acids have been reported as important cofactors promoting human and murine norovirus (NoV) infections in cell culture. The underlying mechanisms are not resolved. Through the use of chemical shift perturbation (CSP) NMR experiments, we identified a low-affinity bile acid binding site of a human GII.4 NoV strain. Long-timescale MD simulations reveal the formation of a ligand-accessible binding pocket of flexible shape, allowing the formation of stable viral coat protein–bile acid complexes in agreement with experimental CSP data. CSP NMR experiments also show that this mode of bile acid binding has a minor influence on the binding of histo-blood group antigens and vice versa. STD NMR experiments probing the binding of bile acids to virus-like particles of seven different strains suggest that low-affinity bile acid binding is a common feature of human NoV and should therefore be important for understanding the role of bile acids as cofactors in NoV infection.     

Mammalian-Like Inflammatory and Pro-Resolving Oxylipins in Marine Algae

Oxylipins constitute a family of oxidized fatty acids, that are well known as tissue hormones in mammals. They contribute to inflammation and its resolution. The major classes of these lipid mediators are inflammatory prostaglandins (PGs) and leukotrienes (LTs) as well as pro-resolving resolvins (Rvs). Understanding their biosynthetic pathways and modes of action is important for anti-inflammatory interventions. Besides mammals, marine algae also biosynthesize mammalian-like oxylipins and thus offer new opportunities for oxylipin research. They provide prolific sources for these compounds and offer unique opportunities to study alternative biosynthetic pathways to the well-known lipid mediators. Herein, we discuss recent findings on the biosynthesis of oxylipins in mammals and algae including an alternative pathway to prostaglandin E₂, a novel pathway to a precursor of leukotriene B₄, and the production of resolvins in algae. We evaluate the pharmacological potential of the algal metabolites with implications in health and disease.     

N-Substituted Nipecotic Acids as (S)-SNAP-5114 Analogues with Modified Lipophilic Domains

Potential mGAT4 inhibitors derived from the lead substance (S)-SNAP-5114 have been synthesized and characterized for their inhibitory potency. Variations from the parent compound included the substitution of one of its aromatic 4-methoxy and 4-methoxyphenyl groups, respectively, with a more polar moiety, including a carboxylic acid, alcohol, nitrile, carboxamide, sulfonamide, aldehyde or ketone function, or amino acid partial structures. Furthermore, it was investigated how the substitution of more than one of the aromatic 4-methoxy groups affects the potency and selectivity of the resulting compounds. Among the synthesized test substances (S)-1-{2-[(4-formylphenyl)bis(4-methoxyphenyl)-methoxy]ethyl}piperidine-3-carboxylic acid, that features a carbaldehyde function in place of one of the aromatic 4-methoxy moieties of (S)-SNAP-5114, was found to have a pIC₅₀ value of 5.89±0.07, hence constituting a slightly more potent mGAT4 inhibitor than the parent substance while showing comparable subtype selectivity.     

Comparison of the wound-activated transformation of caulerpenyne by invasive and noninvasive Caulerpa species of the Mediterranean

The invasive green alga, Caulerpa taxifolia, that has spread rapidly after its introduction into the Mediterranean and the North American Pacific, reacts to wounding by transforming its major metabolite caulerpenyne (1). This wound-activated reaction involves the transformation of the bis-enol acetate moiety of 1, releasing reactive 1,4-dialdehydes. The ability to perform this transformation is found also in both the noninvasive Mediterranean C. prolifera and the invasive C. racemosa. Trapping experiments, as well as transformation of the model substrate geranyl acetate, suggest that all three investigated Caulerpa spp. rely on esterases that act upon wounding of the algae by subsequently removing the three acetate residues of caulerpenyne. The resulting reactive 1,4-dialdehyde oxytoxin 2 (9) can be identified by liquid chromatography–mass spectrometry and is unstable in the wounded tissue. Caulerpenyne transformation occurs rapidly, and severe tissue damage caused degradation of more than 50% of the stored caulerpenyne within 1 min in all three algae. Prevention of the enzymatic reaction before extraction, by shock freezing the tissue with liquid nitrogen, was used for the determination of the caulerpenyne content in intact algae. It gives about twofold higher values compared to an established methanol extraction protocol. The speed and mechanism of the wound-activated transformation, as well as the caulerpenyne content in intact tissue of invasive and noninvasive Caulerpa spp., are comparable. Thus, this enzymatic , transformation, despite being fast and efficient, is likely not the key for the success of the investigated invasive species.     

Influence of dietary fatty acids on the glycerophospholipid composition in organs of cod (gadus morhua)

Cod (mean start weight of 26 g) were fed three diets for 15 months, each based on a dry pellet coated at a level of 9g/100 g with soybean oil, capelin oil or sardine oil. The fatty acid compositions of neutral lipids and four glycerophospholipids of white muscle, liver, gills and heart were determined. The fatty acid composition of dietary lipids influenced the composition of neutral lipids in all organs. Linoleic acid (18∶2n−6) from soybean oil was selectively incorporated into phosphatidylcholine of the four tissues. Similar levels of 20∶5n−3 and 22∶6n−3 in phosphatidylcholine and phosphatidylethanolamine were found in all organs from cod fed capelin oil and sardine oil in spite of highly differentiated feed fatty acid levels. The polyunsaturated fatty acid (PUFA) composition of phosphatidylinositol was least influenced by dietary lipids. The preferred monoenic fatty acid in phospholipids of cod was 18∶1n−9, independent of dietary intake, whereas the longer chain monoenoic acids seemed to be preferentially catabolized. The results suggest that 20∶4n−6 as well as 20∶5n−3 and 22∶6n−3 fatty acids are essential for cod.     

Hyperthermostabilization of Bacillus licheniformis{alpha}-amylase and modulation of its stability over a 50{degrees}C temperature range

Bacillus licheniformis