On-line (HPLC-NMR) and off-line phytochemical profiling of the Australian plant, Lasiopetalum macrophyllum

On-line (HPLC-NMR) and off-line (HPLC, NMR and MS) methodologies were used to profile the constituents present in the crude extract of Lasiopetalum macrophyllum. On-flow and stop-flow HPLC-NMR supported the presence of trans-tiliroside and permitted partial identification of cis-tiliroside and 4'-methoxy-trans-tiliroside. Off-line isolation led to the unequivocal identification of four flavanoid glycosides including a new structural derivative, 4'-methoxy-cis-tiliroside. This is the first report of flavonoid glycosides occurring in this plant genus. In addition, a number of structure revisions have been proposed for previously reported flavonoid glycosides that were incorrectly assigned

Muscular diacylglycerol metabolism and insulin resistance

Failure of insulin to elicit an increase in glucose uptake and metabolism in target tissues such as skeletal muscle is a major characteristic of non-insulin dependent type 2 diabetes mellitus. A strong correlation between intramyocellular triacylglycerol concentrations and the severity of insulin resistance has been found and led to the assumption that lipid oversupply to skeletal muscle contributes to reduced insulin action. However, the molecular mechanism that links intramyocellular lipid content with the generation of muscle insulin resistance is still unclear. It appears unlikely that the neutral lipid metabolite triacylglycerol directly impairs insulin action. Hence it is believed that intermediates in fatty acid metabolism, such as fatty acyl-CoA, ceramides or diacylglycerol (DAG) link fat deposition in the muscle to compromised insulin signaling. DAG is identified as a potential mediator of lipid-induced insulin resistance, as increased DAG levels are associated with protein kinase C activation and a reduction in both insulin-stimulated IRS-1 tyrosine phosphorylation and PI3 kinase activity. As DAG is an intermediate in the synthesis of triacylglycerol from fatty acids and glycerol, its level can be lowered by either improving the oxidation of cellular fatty acids or by accelerating the incorporation of fatty acids into triacylglycerol. This review discusses the evidence that implicates DAG being central in the development of muscular insulin resistance. Furthermore, we will discuss if and how modulation of skeletal muscle DAG levels could function as a possible therapeutic target for the treatment of type 2 diabetes mellitus

Dynamic control of selectivity in the ubiquitination pathway revealed by an ASP to GLU substitution in an intra-molecular salt-bridge network

Ubiquitination relies on a subtle balance between selectivity and promiscuity achieved through specific interactions between ubiquitin-conjugating enzymes (E2s) and ubiquitin ligases (E3s). Here, we report how a single aspartic to glutamic acid substitution acts as a dynamic switch to tip the selectivity balance of human E2s for interaction toward E3 RING-finger domains. By combining molecular dynamic simulations, experimental yeast-two-hybrid screen of E2-E3 (RING) interactions and mutagenesis, we reveal how the dynamics of an internal salt-bridge network at the rim of the E2-E3 interaction surface controls the balance between an “open”, binding competent, and a “closed”, binding incompetent state. The molecular dynamic simulations shed light on the fine mechanism of this molecular switch and allowed us to identify its components, namely an aspartate/glutamate pair, a lysine acting as the central switch and a remote aspartate. Perturbations of single residues in this network, both inside and outside the interaction surface, are sufficient to switch the global E2 interaction selectivity as demonstrated experimentally. Taken together, our results indicate a new mechanism to control E2-E3 interaction selectivity at an atomic level, highlighting how minimal changes in amino acid side-chain affecting the dynamics of intramolecular salt-bridges can be crucial for protein-protein interactions. These findings indicate that the widely accepted sequence-structure-function paradigm should be extended to sequence-structure-dynamics-function relationship and open new possibilities for control and fine-tuning of protein interaction selectivity

Disentangling conical intersection and coherent molecular dynamics in methyl bromide with attosecond transient absorption spectroscopy

Attosecond probing of core-level electronic transitions provides a sensitive tool for studying valence molecular dynamics with atomic, state, and charge specificity. In this report, we employ attosecond transient absorption spectroscopy to follow the valence dynamics of strong-field initiated processes in methyl bromide. By probing the 3d core-to-valence transition, we resolve the strong field excitation and ensuing fragmentation of the neutral σ* excited states of methyl bromide. The results provide a clear signature of the non-adiabatic passage of the excited state wavepacket through a conical intersection. We additionally observe competing, strong field initiated processes arising in both the ground state and ionized molecule corresponding to vibrational and spin-orbit motion, respectively. The demonstrated ability to resolve simultaneous dynamics with few-femtosecond resolution presents a clear path forward in the implementation of attosecond XUV spectroscopy as a general tool for probing competing and complex molecular phenomena with unmatched temporal resolution

Metabolism and occurrence of methanogenic and sulfate-reducing syntrophic acetate oxidizing communities in haloalkaline environments

The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fmicb. 2018.03039/full#supplementary-materialAnaerobic syntrophic acetate oxidation (SAO) is a thermodynamically unfavorable process involving a syntrophic acetate oxidizing bacterium (SAOB) that forms interspecies electron carriers (IECs). These IECs are consumed by syntrophic partners, typically hydrogenotrophic methanogenic archaea or sulfate reducing bacteria. In this work, the metabolism and occurrence of SAOB at extremely haloalkaline conditions were investigated, using highly enriched methanogenic (M-SAO) and sulfate-reducing (S-SAO) cultures from south-eastern Siberian hypersaline soda lakes. Activity tests with the M-SAO and S-SAO cultures and thermodynamic calculations indicated that hydrogen and formate are important IECs in both SAO cultures. Metagenomic analysis of the M-SAO cultures showed that the dominant SAOB was Candidatus Syntrophonatronum acetioxidans, and a near-complete draft genome of this SAOB was reconstructed. Ca. S. acetioxidans has all genes necessary for operating the Wood-Ljungdahl pathway, which is likely employed for acetate oxidation. It also encodes several genes essential to thrive at haloalkaline conditions; including a Na+-dependent ATP synthase and marker genes for salt-out strategies for osmotic homeostasis at high soda conditions. Membrane lipid analysis of the M-SAO culture showed the presence of unusual bacterial diether membrane lipids which are presumably beneficial at extreme haloalkaline conditions. To determine the importance of SAO in haloalkaline environments, previously obtained 16S rRNA gene sequencing data and metagenomic data of five different hypersaline soda lake sediment samples were investigated, including the soda lakes where the enrichment cultures originated from. The draft genome of Ca. S. acetioxidans showed highest identity with two metagenome-assembled genomes (MAGs) of putative SAOBs that belonged to the highly abundant and diverse Syntrophomonadaceae family present in the soda lake sediments. The 16S rRNA amplicon datasets of the soda lake sediments showed a high similarity of reads to Ca. S. acetioxidans with abundance as high as 1.3% of all reads, whereas aceticlastic methanogens and acetate oxidizing sulfate-reducers were not abundant (0.1%) or could not be detected. These combined results indicate that SAO is the primary anaerobic acetate oxidizing pathway at extreme haloalkaline conditions performed by haloalkaliphilic syntrophic consortia.This research was supported by the Soehngen Institute of AnaerobicMicrobiology(SIAM) Gravitation grant(024.002.002) of the Netherlands Ministry of Education, Culture and Science and the Netherlands Organisation for Scientific Research (NWO). GM and CV were supported by the ERC Advanced Grant PARASOL (No. 322551). DS also received support from the Russian Foundation for Basic Research (16-04-00035) and the Russian Academy of Sciences and Federal Agency of Scientific Organizations(0104-2018-0033), AS by the ERC Advanced Grant Novel Anaerobes (No. 323009), and JD by the ERC Advanced Grant Microlipids (No.694569).info:eu-repo/semantics/publishedVersio