Approach to the homoerythrina alkaloids using a tandem N-alkylation/azomethine ylide cycloaddition

Synthetic efforts toward the homoerythrina alkaloids 1-3 are described. Two separate model systems guided the pivotal [3 + 2] azomethine ylide cycloaddition cascade to form the A-C rings of these alkaloids. The cycloaddition precursors 63 and 68, prepared in nine and ten steps, respectively, from alkyne 47, each contain an enolizable ketone, a tethered electrophile, and an electron-poor dipolarophile. Heating 63 and 68 with the stannyl amine 17 generated demethoxyschelhammeridine 65 and demethoxyschelhammericine 70, the products of intramolecular azomethine ylide cycloadditions. Subsequent attempts to install the C-3 methoxy group of 1-3 are also described.     

Optimization of unnatural base pair packing for polymerase recognition

As part of an effort to expand the genetic alphabet, we have been examining the ability of predominately hydrophobic nucleobase analogues to pair in duplex DNA and during polymerase-mediated replication. We previously reported the synthesis and thermal stability of unnatural base pairs formed between nucleotides bearing simple methyl-substituted phenyl ring nucleobase analogues. Several of these pairs are virtually as stable and selective as natural base pairs in the same sequence context. Here, we report the characterization of polymerase-mediated replication of the same unnatural base pairs. We find that every facet of replication, including correct and incorrect base pair synthesis, as well as continued primer extension beyond the unnatural base pair, is sensitive to the specific methyl substitution pattern of the nucleobase analogue. The results demonstrate that neither hydrogen bonding nor large aromatic surface area is required for polymerase recognition, and that interstrand interactions between small aromatic rings may be optimized for replication. Combined with our previous results, these studies suggest that appropriately derivatized phenyl nucleobase analogues represent a promising approach toward developing a third base pair and expanding the genetic alphabet.     

Alkali-metal-mediated zincation of anisole: synthesis and structures of three instructive ortho-zincated complexes

The new concept of alkali-metal-mediated zincation (AMMZ), formally a zinc-hydrogen exchange reaction but one that requires the participation of an alkali metal, is applied here to the alkyl aryl ether anisole, an important molecule for studying directed ortho-metalation (DoM) chemistry. Treating one molar equivalent of anisole with the lithium dialkyl-TMP zincate reagent [THF.Li(mu-TMP)(mu-tBu)Zn(tBu)] (1) in hexane solution affords the mono-ortho-zincated complex [THF.Li(mu-TMP)(mu-o-C6H4OMe)Zn(tBu)] (2), which establishes that 1 functions as an alkyl base although previously it was regarded as an amido (TMP) base in other DoM applications. Treating two molar equivalents of anisole with 1, and increasing the reaction time, affords the bis-ortho-zincated complex [THF.Li(mu-TMP)(mu-o-C6H4OMe)Zn(o-C6H4OMe)] (3), which establishes that 1 can also function as a dual alkyl base. Omitting THF and rerunning the reaction with one or two molar equivalents of anisole affords [Ph(Me)O.Li(mu-TMP)(mu-o-C6H4OMe)Zn(tBu)] (4), which remarkably contains a combination of neutral and ortho-deprotonated anisole ligands. On isolating crystalline 4 from solution and adding THF, it converts to 2 and then to 3 on further stirring of the solution, as determined by NMR studies. This fact, along with other observations, would suggest that a complex-induced proximity effect does not need to be invoked to explain the observed zincation of anisole. The crystal structures of 2-4 are presented, as are their 1H, 13C, and 7Li NMR spectra recorded in C6D6 solution.     

Negative Ions Enhance Survival of Membrane Protein Complexes

Membrane protein complexes are commonly introduced to the mass spectrometer solubilized in detergent micelles. The collisional activation used to remove the detergent, however, often causes protein unfolding and dissociation. As in the case for soluble proteins, electrospray in the positive ion mode is most commonly used for the study of membrane proteins. Here we show several distinct advantages of employing the negative ion mode. Negative polarity can yield lower average charge states for membrane proteins solubilized in saccharide detergents, with enhanced peak resolution and reduced adduct formation. Most importantly, we demonstrate that negative ion mode electrospray ionization (ESI) minimizes subunit dissociation in the gas phase, allowing access to biologically relevant oligomeric states. Together, these properties mean that intact membrane protein ions can be generated in a greater range of solubilizing detergents. The formation of negative ions, therefore, greatly expands the possibilities of using mass spectrometry on this intractable class of protein.

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On developing coarse-grained models for biomolecular simulation: a review

So-called coarse-grained models are a popular type of model for accessing long time scales in simulations of biomolecular processes. Such models are coarse-grained with respect to atomic models. But any modelling of processes or substances involves coarse-graining, i.e. the elimination of non-essential degrees of freedom and interactions from a more fine-grained level of modelling. The basic ingredients of developing coarse-grained models based on the properties of fine-grained models are reviewed, together with the conditions that must be satisfied in order to preserve the correct physical mechanisms in the coarse-graining process. This overview should help the reader to determine how realistic a coarse-grained model of a biomolecular system is, i.e. whether it reflects the underlying physical mechanisms or merely provides a set of pretty pictures of the process or substances of interest.     

Zinc(II) complexes of constrained antiviral macrocycles

The configurations of metallocyclams are of interest in relation to protein recognition and anti-HIV activity. We have synthesised four novel zinc(II) complexes with hexyl-Me(2)-cyclam (HMC; 3,14-dimethyl-2,6,13,17-tetraazatricyclo(16.4.0.0(7,12))docosane), 1, and naphthyl-hexyl-Me(2)-cyclam (NHMC; 2,13-bis(1-naphthylmethyl)-5,16-dimethyl-2,6,13,17-tetraazatricyclo(16.4.0.0(7,12))docosane), 2, as ligands. X-ray crystallographic data for Zn(II)-HMC diacetate, 3 show that zinc is six-coordinate in a distorted octahedral environment bound to four equatorial N atoms from the macrocycle and two axial acetato O atoms. The 14-membered metallo-macrocycle adopts a trans-III (RRSS) configuration with two six-membered rings in chair forms and two five-membered rings in gauche forms. In the chlorido Zn(II)-HMC complex 5, zinc appears to be 5-coordinate with square-pyramidal geometry. Interestingly, the chlorido Zn(II)-NHMC complex 6 crystallised in a trans-I configuration containing 4-coordinate tetrahedral zinc bound to three cyclam ring N atoms, a possible model for intermediates formed during the uptake and release of metals by cyclams. The ligand 1 and the zinc complex 3 were active towards viral strains HIV-1 (III(B)) (IC(50) values of 10.51 ± 0.23 and 3.50 ± 0.33 μM, respectively), and HIV-2 (ROD) (IC(50) values of 133.78 ± 14.10 and >110.67 μM, respectively). 2D [(1)H, (13)C] and [(1)H, (15)N] NMR spectroscopic studies suggested that the types of configurational isomers present in solution depend on the axial ligand.     

A microfluidic spiral for size-dependent fractionation of magnetic microspheres

Magnetic microspheres (MMS) are useful tools for a variety of medical and pharmaceutical applications. Typically, commercially manufactured MMS exhibit broad size distributions. This polydispersity is problematic for many applications. Since the direct synthesis of monodisperse MMS is often fraught with technical challenges, there is considerable interest in and need associated with the development of techniques for size-dependent fractionation of MMS. In this study we demonstrated continuous size-dependent fractionation of sub-micron scale particles driven by secondary (Dean effect) flows in curved microfluidic channels. Our goal was to demonstrate that such techniques can be applied to MMS containing superparamagnetic nanoparticles. To achieve this goal, we developed and tested a microfluidic chip for continuous MMS fractionation. Our data address two key areas. First, the densities of MMS are typically in the range 1.5–2.5 g/cm³, and thus they tend be non-neutrally buoyant. Our data demonstrate that efficient size-dependent fractionation of MMS entrained in water (density 1 g/cm³) is possible and is not significantly influenced by the density mismatch. In this context we show that a mixture comprising two different monodisperse MMS components can be separated into its constituent parts with 100% and 88% success for the larger and smaller particles, respectively. Similarly, we show that a suspension of polydisperse MMS can be separated into streams containing particles with different mean diameters. Second, our data demonstrate that efficient size-dependent fractionation of MMS is not impeded by magnetic interactions between particles, even under application of homogeneous magnetic fields as large as 35 kA/m. The chip is thus suitable for the separation of different particle fractions in a continuous process and the size fractions can be chosen simply by adjusting the flow velocity of the carrier fluid. These facts open the door to size dependent fractionation of MMS.     

Nanoengineered glycan sensors enabling native glycoprofiling for medicinal applications: towards profiling glycoproteins without labeling or liberation steps

Nanoengineered glycan sensors may help realize the long-held goal of accurate and rapid glycoprotein profiling without labeling or glycan liberation steps. Current methods of profiling oligosaccharides displayed on protein surfaces, such as liquid chromatography, mass spectrometry, capillary electrophoresis, and microarray methods, are limited by sample pretreatment and quantitative accuracy. Microarrayed platforms can be improved with methods that better estimate kinetic parameters rather than simply reporting relative binding information. These quantitative glycan sensors are enabled by an emerging class of nanoengineered materials that differ in their mode of signal transduction from traditional methods. Platforms that respond to mass changes include a quartz crystal microbalance and cantilever sensors. Electronic response can be detected from electrochemical, field effect transistor, and pore impedance sensors. Optical methods include fluorescent frontal affinity chromatography, surface plasmon resonance methods, and fluorescent carbon nanotubes. After a very brief primer on glycobiology and its connection to medicine, these emerging systems are critically reviewed for their potential use as core sensors in future glycoprofiling tools.     

Reduced‐order modelling of an adaptive mesh ocean model

A novel proper orthogonal decomposition (POD) model has been developed for use with an advanced unstructured mesh finite‐element ocean model, the Imperial College Ocean Model (ICOM, described in detail below), which includes many recent developments in ocean modelling and numerical analysis. The advantages of the POD model developed here over existing POD approaches are the ability:

• 1. To increase accuracy when representing geostrophic balance (the balance between the Coriolis terms and the pressure gradient). This is achieved through the use of two sets of geostrophic basis functions where each one is calculated by basis functions for velocities u and v.
• 2. To speed up the POD simulation. To achieve this a new numerical technique is introduced, whereby a time‐dependent matrix in the discretized equation is rapidly constructed from a series of time‐independent matrices. This development imparts considerable efficiency gains over the often‐used alternative of calculating each finite element over the computational domain at each time level.
• 3. To use dynamically adaptive meshes in the above POD model.

Copyright © 2008 John Wiley & Sons, Ltd.