Bacterial biofilms are cellular communities that produce an adherent matrix. Exopolysaccharides are key structural components of this matrix and are required for the assembly and architecture of biofilms produced by a wide variety of microorganisms. The human bacterial pathogens Escherichia coli and Salmonella enterica produce a biofilm matrix composed primarily of the exopolysaccharide phosphoethanolamine (pEtN) cellulose. Once thought to be composed of only underivatized cellulose, the pEtN modification present in these matrices has been implicated in the overall architecture and integrity of the biofilm. However, an understanding of the mechanism underlying pEtN derivatization of the cellulose exopolysaccharide remains elusive. The bacterial cellulose synthase subunit G (BcsG) is a predicted inner membrane–localized metalloenzyme that has been proposed to catalyze the transfer of the pEtN group from membrane phospholipids to cellulose. Here we present evidence that the C-terminal domain of BcsG from E. coli (EcBcsGΔN) functions as a phosphoethanolamine transferase in vitro with substrate preference for cellulosic materials. Structural characterization of EcBcsGΔN revealed that it belongs to the alkaline phosphatase superfamily, contains a Zn2+ ion at its active center, and is structurally similar to characterized enzymes that confer colistin resistance in Gram-negative bacteria. Informed by our structural studies, we present a functional complementation experiment in E. coli AR3110, indicating that the activity of the BcsG C-terminal domain is essential for integrity of the pellicular biofilm. Furthermore, our results established a similar but distinct active-site architecture and catalytic mechanism shared between BcsG and the colistin resistance enzymes.

8 January 2020 , Volume 96, Number 1

4 March 2020 , Volume 96, Number 2

Purinergic signaling by extracellular ATP regulates a variety of cellular events and is implicated in both normal physiology and pathophysiology. Several molecules have been associated with the release of ATP and other small molecules, but their precise contributions have been difficult to assess because of their complexity and heterogeneity. Here, we report on the results of a gain-of-function screen for modulators of hypotonicity-induced ATP release using HEK-293 cells and murine cerebellar granule neurons, along with bioluminescence, calcium FLIPR, and short hairpin RNA–based gene-silencing assays. This screen utilized the most extensive genome-wide ORF collection to date, covering 90% of human, nonredundant, protein-encoding genes. We identified two ABCG1 (ABC subfamily G member 1) variants, which regulate cellular cholesterol, as modulators of hypotonicity-induced ATP release. We found that cholesterol levels control volume-regulated anion channel–dependent ATP release. These findings reveal novel mechanisms for the regulation of ATP release and volume-regulated anion channel activity and provide critical links among cellular status, cholesterol, and purinergic signaling.

Autoimmune thyroid diseases (AITD) are the most common group of autoimmune diseases, associated with lymphocyte infiltration and the production of thyroid autoantibodies, like thyroid peroxidase antibodies (TPOAb), in the thyroid gland. Immunoglobulins and cell-surface receptors are glycoproteins with distinctive glycosylation patterns that play a structural role in maintaining and modulating their functions. We investigated associations of total circulating IgG and peripheral blood mononuclear cells glycosylation with AITD and the influence of genetic background in a case-control study with several independent cohorts and over 3,000 individuals in total. The study revealed an inverse association of IgG core fucosylation with TPOAb and AITD, as well as decreased peripheral blood mononuclear cells antennary α1,2 fucosylation in AITD, but no shared genetic variance between AITD and glycosylation. These data suggest that the decreased level of IgG core fucosylation is a risk factor for AITD that promotes antibody-dependent cell-mediated cytotoxicity previously associated with TPOAb levels.

The study of protein subcellular distribution, their assembly into complexes and the set of proteins with which they interact with is essential to our understanding of fundamental biological processes. Complementary to traditional assays, proximity-dependent biotinylation (PDB) approaches coupled with mass spectrometry (such as BioID or APEX) have emerged as powerful techniques to study proximal protein interactions and the subcellular proteome in the context of living cells and organisms. Since their introduction in 2012, PDB approaches have been used in an increasing number of studies and the enzymes themselves have been subjected to intensive optimization. How these enzymes have been optimized and considerations for their use in proteomics experiments are important questions. Here, we review the structural diversity and mechanisms of the two main classes of PDB enzymes: the biotin protein ligases (BioID) and the peroxidases (APEX). We describe the engineering of these enzymes for PDB and review emerging applications, including the development of PDB for coincidence detection (split-PDB). Lastly, we briefly review enzyme selection and experimental design guidelines and reflect on the labeling chemistries and their implication for data interpretation.

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Publication Date: Apr 10 2020 The SEC will withdraw the existing approval order and the fund will become subject to the requirements of Rule 6c-11 and Nasdaq Rule 5704....

Publication Date: Apr 10 2020 On or before December 22, 2020, all ETFs that meet the definition of "Exchange Traded Fund" in Nasdaq Rule 5704(a)(1)(A) will need to be compliant with Rule 6c-11. If it is determined that an ETF no longer complies with Rule 6c-11 and therefore no longer complies Nasdaq Rule 5704, Nasdaq will generally issue a letter of deficiency. The ETF will generally be given 45 days to submit a plan to regain compliance. If the plan is accepted, Nasdaq Staff can grant an...

8 January 2020 , Volume 96, Number 1

The music of most hit songs is pretty well known, but sometimes there are mysteries. One question that remained unanswered for over forty years is: What instrumentation and notes make up the opening chord of the Beatles. "A Hard Day.s Night"? Mathematician Jason Brown - a big Beatles fan - recently solved the puzzle using his musical knowledge and discrete Fourier transforms, mathematical transformations that help decompose signals into their basic parts. These transformations simplify applications ranging from signal processing to multiplying large numbers, so that a researcher doesn.t have to be "working like a dog" to get an answer. Brown is also using mathematics, specifically graph theory, to discover who wrote "In My Life," which both Lennon and McCartney claimed to have written. In his graphs, chords are represented by points that are connected when one chord immediately follows another. When all songs with known authorship are diagrammed, Brown will see which collection of graphs - McCartney.s or Lennon.s - is a better fit for "In My Life." Although it may seem a bit counterintuitive to use mathematics to learn more about a revolutionary band, these analytical methods identify and uncover compositional principles inherent in some of the best Beatles. music. Thus it.s completely natural and rewarding to apply mathematics to the Fab 4 For More Information: Professor Uses Mathematics to Decode Beatles Tunes, "The Wall Street Journal", January 30, 2009..

David Murray-Hundley, our resident Grumpy Entrepreneur, on the eight things for startups to consider if the bubble bursts