A general catalytic β-C-H carbonylation of aliphatic amines to β-lactams

Methods for the synthesis and functionalization of amines are intrinsically important to a variety of chemical applications. We present a general carbon-hydrogen bond activation process that combines readily available aliphatic amines and the feedstock gas carbon monoxide to form synthetically versatile value-added amide products. The operationally straightforward palladium-catalyzed process exploits a distinct reaction pathway, wherein a sterically hindered carboxylate ligand orchestrates an amine attack on a palladium anhydride to transform aliphatic amines into β-lactams. The reaction is successful with a wide range of secondary amines and can be used as a late-stage functionalization tactic to deliver advanced, highly functionalized amine products of utility for pharmaceutical research and other areas.We gratefully acknowledge funding from the European Research Council and the UK Engineering and Physical Sciences Research Council (EPSRC) (to D.W., K.F.H., A.P.S., and M.J.G.), the Herchel Smith Trust (to B.G.N.C.), the Marie Curie Foundation (to J.C.), and the Royal Society (Wolfson Merit Award to M.J.G.). Mass spectrometry data were acquired at the EPSRC UK National Mass Spectrometry Facility at Swansea University. Computational work was performed with the Darwin Supercomputer of the University of Cambridge High Performance Computing Service (http://www.hpc.cam.ac.uk/), provided by Dell, using Strategic Research Infrastructure Funding from the Higher Education Funding Council for England

Tuning the Anthranilamide Peptidomimetic Design to Selectively Target Planktonic Bacteria and Biofilm

There is a pressing need to develop new antimicrobials to help combat the increase in antibiotic resistance that is occurring worldwide. In the current research, short amphiphilic antibacterial and antibiofilm agents were produced by tuning the hydrophobic and cationic groups of anthranilamide peptidomimetics. The attachment of a lysine cationic group at the tail position increased activity against E. coli by >16-fold (from >125 μM to 15.6 μM) and greatly reduced cytotoxicity against mammalian cells (from ≤20 μM to ≥150 μM). These compounds showed significant disruption of preformed biofilms of S. aureus at micromolar concentrations

Matched sizes of activating and inhibitory receptor/ligand pairs are required for optimal signal integration by human Natural Killer cells

It has been suggested that receptor-ligand complexes segregate or co-localise within immune synapses according to their size, and this is important for receptor signaling. Here, we set out to test the importance of receptor-ligand complex dimensions for immune surveillance of target cells by human Natural Killer (NK) cells. NK cell activation is regulated by integrating signals from activating receptors, such as NKG2D, and inhibitory receptors, such as KIR2DL1. Elongating the NKG2D ligand MICA reduced its ability to trigger NK cell activation. Conversely, elongation of KIR2DL1 ligand HLA-C reduced its ability to inhibit NK cells. Whereas normal-sized HLA-C was most effective at inhibiting activation by normal-length MICA, only elongated HLA-C could inhibit activation by elongated MICA. Moreover, HLA-C and MICA that were matched in size co-localised, whereas HLA-C and MICA that were different in size were segregated. These results demonstrate that receptor-ligand dimensions are important in NK cell recognition, and suggest that optimal integration of activating and inhibitory receptor signals requires the receptor-ligand complexes to have similar dimensions

Serum albumin and osmolality inhibit Bdellovibrio bacteriovorus predation in human serum

We evaluated the bactericidal activity of Bdellovibrio bacteriovorus, strain HD100, within blood sera against bacterial strains commonly associated with bacteremic infections, including E. coli, Klebsiella pneumoniae and Salmonella enterica. Tests show that B. bacteriovorus HD100 is not susceptible to serum complement or its bactericidal activity. After a two hour exposure to human sera, the prey populations decreased 15- to 7,300-fold due to the serum complement activity while, in contrast, the B. bacteriovorus HD100 population showed a loss of only 33%. Dot blot analyses showed that this is not due to the absence of antibodies against this predator. Predation in human serum was inhibited, though, by both the osmolality and serum albumin. The activity of B. bacteriovorus HD100 showed a sharp transition between 200 and 250 mOsm/kg, and was progressively reduced as the osmolality increased. Serum albumin also acted to inhibit predation by binding to and coating the predatory cells. This was confirmed via dot blot analyses and confocal microscopy. The results from both the osmolality and serum albumin tests were incorporated into a numerical model describing bacterial predation of pathogens. In conclusion, both of these factors inhibit predation and, as such, they limit its effectiveness against pathogenic prey located within sera

TGF-ß Sma/Mab Signaling Mutations Uncouple Reproductive Aging from Somatic Aging

Female reproductive cessation is one of the earliest age-related declines humans experience, occurring in mid-adulthood. Similarly, Caenorhabditis elegans' reproductive span is short relative to its total life span, with reproduction ceasing about a third into its 15–20 day adulthood. All of the known mutations and treatments that extend C. elegans' reproductive period also regulate longevity, suggesting that reproductive span is normally linked to life span. C. elegans has two canonical TGF-ß signaling pathways. We recently found that the TGF-ß Dauer pathway regulates longevity through the Insulin/IGF-1 Signaling (IIS) pathway; here we show that this pathway has a moderate effect on reproductive span. By contrast, TGF-ß Sma/Mab signaling mutants exhibit a substantially extended reproductive period, more than doubling reproductive span in some cases. Sma/Mab mutations extend reproductive span disproportionately to life span and act independently of known regulators of somatic aging, such as Insulin/IGF-1 Signaling and Dietary Restriction. This is the first discovery of a pathway that regulates reproductive span independently of longevity and the first identification of the TGF-ß Sma/Mab pathway as a regulator of reproductive aging. Our results suggest that longevity and reproductive span regulation can be uncoupled, although they appear to normally be linked through regulatory pathways

Environmental Control of Phase Transition and Polyp Survival of a Massive-Outbreaker Jellyfish

A number of causes have been proposed to account for the occurrence of gelatinous zooplankton (both jellyfish and ctenophore) blooms. Jellyfish species have a complex life history involving a benthic asexual phase (polyp) and a pelagic sexual phase (medusa). Strong environmental control of jellyfish life cycles is suspected, but not fully understood. This study presents a comprehensive analysis on the physicochemical conditions that control the survival and phase transition of Cotylorhiza tuberculata; a scyphozoan that generates large outbreaks in the Mediterranean Sea. Laboratory experiments indicated that the influence of temperature on strobilation and polyp survival was the critical factor controlling the capacity of this species to proliferate. Early life stages were less sensitive to other factors such as salinity variations or the competitive advantage provided by zooxanthellae in a context of coastal eutrophication. Coherently with laboratory results, the presence/absence of outbreaks of this jellyfish in a particular year seems to be driven by temperature. This is the first time the environmental forcing of the mechanism driving the life cycle of a jellyfish has been disentangled via laboratory experimentation. Projecting this understanding to a field population under climatological variability results in a pattern coherent with in situ records

An Eye to a Kill: Using Predatory Bacteria to Control Gram-Negative Pathogens Associated with Ocular Infections

Ocular infections are a leading cause of vision loss. It has been previously suggested that predatory prokaryotes might be used as live antibiotics to control infections. In this study, Pseudomonas aeruginosa and Serratia marcescens ocular isolates were exposed to the predatory bacteria Micavibrio aeruginosavorus and Bdellovibrio bacteriovorus. All tested S. marcescens isolates were susceptible to predation by B. bacteriovorus strains 109J and HD100. Seven of the 10 P. aeruginosa isolates were susceptible to predation by B. bacteriovorus 109J with 80% being attacked by M. aeruginosavorus. All of the 19 tested isolates were found to be sensitive to at least one predator. To further investigate the effect of the predators on eukaryotic cells, human corneal-limbal epithelial (HCLE) cells were exposed to high concentrations of the predators. Cytotoxicity assays demonstrated that predatory bacteria do not damage ocular surface cells in vitro whereas the P. aeruginosa used as a positive control was highly toxic. Furthermore, no increase in the production of the proinflammatory cytokines IL-8 and TNF-alpha was measured in HCLE cells after exposure to the predators. Finally, injection of high concentration of predatory bacteria into the hemocoel of Galleria mellonella, an established model system used to study microbial pathogenesis, did not result in any measurable negative effect to the host. Our results suggest that predatory bacteria could be considered in the near future as a safe topical bio-control agent to treat ocular infections. © 2013 Shanks et al

High Affinity Antigen Recognition of the Dual Specific Variants of Herceptin Is Entropy-Driven in Spite of Structural Plasticity

The antigen-binding site of Herceptin, an anti-human Epidermal Growth Factor Receptor 2 (HER2) antibody, was engineered to add a second specificity toward Vascular Endothelial Growth Factor (VEGF) to create a high affinity two-in-one antibody bH1. Crystal structures of bH1 in complex with either antigen showed that, in comparison to Herceptin, this antibody exhibited greater conformational variability, also called “structural plasticity”. Here, we analyzed the biophysical and thermodynamic properties of the dual specific variants of Herceptin to understand how a single antibody binds two unrelated protein antigens. We showed that while bH1 and the affinity-improved bH1-44, in particular, maintained many properties of Herceptin including binding affinity, kinetics and the use of residues for antigen recognition, they differed in the binding thermodynamics. The interactions of bH1 and its variants with both antigens were characterized by large favorable entropy changes whereas the Herceptin/HER2 interaction involved a large favorable enthalpy change. By dissecting the total entropy change and the energy barrier for dual interaction, we determined that the significant structural plasticity of the bH1 antibodies demanded by the dual specificity did not translate into the expected increase of entropic penalty relative to Herceptin. Clearly, dual antigen recognition of the Herceptin variants involves divergent antibody conformations of nearly equivalent energetic states. Hence, increasing the structural plasticity of an antigen-binding site without increasing the entropic cost may play a role for antibodies to evolve multi-specificity. Our report represents the first comprehensive biophysical analysis of a high affinity dual specific antibody binding two unrelated protein antigens, furthering our understanding of the thermodynamics that drive the vast antigen recognition capacity of the antibody repertoire