Quantification of Structural Integrity and Stability Using Nanograms of Protein by Flow-Induced Dispersion Analysis

In the development of therapeutic proteins, analytical assessment of structural stability and integrity constitutes an important activity, as protein stability and integrity influence drug efficacy, and ultimately patient safety. Existing analytical methodologies solely rely on relative changes in optical properties such as fluorescence or scattering upon thermal or chemical perturbation. Here, we present an absolute analytical method for assessing protein stability, structure, and unfolding utilizing Taylor dispersion analysis (TDA) and LED-UV fluorescence detection. The developed TDA method measures the change in size (hydrodynamic radius) and intrinsic fluorescence of a protein during in-line denaturation with guanidinium hydrochloride (GuHCl). The conformational stability of the therapeutic antibody adalimumab and human serum albumin were characterized as a function of pH. The simple workflow and low sample consumption (40 ng protein per data point) of the methodology make it ideal for assessing protein characteristics related to stability in early drug development or when having a scarce amount of sample available.     

Shell filling in closed single-wall carbon nanotube quantum dots

We observe twofold shell filling in the spectra of closed one-dimensional quantum dots formed in single-wall carbon nanotubes. Its signatures include a bimodal distribution of addition energies, correlations in the excitation spectra for different electron number, and alternation of the spins of the added electrons. This provides a contrast with quantum dots in higher dimensions, where such spin pairing is absent. We also see indications of an additional fourfold periodicity indicative of K-K' subband shells. Our results suggest that the absence of shell filling in most isolated nanotube dots results from disorder or nonuniformity.     

Enantioselective synthetic thalidomide receptors based upon DNA binding motifs

A series of molecularly imprinted polymer (MIP) synthetic receptors selective for the sedative thalidomide (5) have been designed and synthesized based upon the functional monomer 9-(2'-methacryloyloxyethyl)adenine (2). (1)H-NMR studies were used to establish the existence of DNA-like binding interactions between 2 and the template (5). A series of ethylene glycol dimethacrylate cross-linked copolymers was synthesized using either 2 or methacrylic acid, or a combination of these functional monomers. Zonal HPLC studies demonstrated enantioselectivity (alpha = 2.11) and ligand selectivity which could be attributed to the interaction of 2 with the imide moiety of 5. Compound 2 provided a more significant contribution to the binding of 5 than methacrylic acid, though a combination of these two functional monomers resulted in improved enantioselectivity. Frontal chromatographic and batch binding studies confirmed the observed differences in affinity of the imprinted and reference polymers for the template.     

Semi-Balanced Colorings of Graphs: Generalized 2-Colorings Based on a Relaxed Discrepancy Condition

We generalize the concept of a 2-coloring of a graph to what we call a semi-balanced coloring by relaxing a certain discrepancy condition on the shortest-paths hypergraph of the graph. Let G be an undirected, unweighted, connected graph with n vertices and m edges. We prove that the number of different semi-balanced colorings of G is: (1) at most n+1 if G is bipartite; (2) at most m if G is non-bipartite and triangle-free; and (3) at most m+1 if G is non-bipartite. Based on the above combinatorial investigation, we design an algorithm to enumerate all semi-balanced colorings of G in O(nm²) time.Acknowledgments The authors thank Tetsuo Asano, Naoki Katoh, Kunihiko Sadakane, and Hisao Tamaki for helpful discussions and comments.Supported in part by Sweden-Japan FoundationFinal version received: November 17, 2003     

Synthesis of pentacyclic 13-azadibenzo[a,de]anthracenes via anionic cascade ring closure

Bromine-lithium exchange using tert-butyllithium at -78 degrees C initiates a cascade process whereby either xanthone derivatives or pentacyclic 13-azadibenzo[a,de]anthracenes are produced in high yields. The reaction proceeds via a sequential intramolecular trapping of organolithium intermediates.     

Monosaccharide templates for de novo designed 4-alpha-helix bundle proteins: template effects in carboproteins

De novo design and total chemical synthesis of proteins provide powerful approaches to critically test our understanding of protein folding, structure, and stability. The 4-alpha-helix bundle is a frequently studied structure in which four amphiphilic alpha-helical peptide strands form a hydrophobic core. Assembly of protein models on a template has been suggested as a way to reduce the entropy of folding. We have previously developed the concept of carbohydrates as templates in the de novo design of protein models termed 'carboproteins'. Here we present the chemical synthesis of three 8.1 kDa 4-alpha-helix bundles by oxime ligation of tetra-aminooxyacetyl functionalized D-galacto-, D-gluco-, and D-altropyranoside templates with an amphiphilic C-terminal hexadecapeptide aldehyde sequence. CD spectroscopy indicated that the choice of template has an effect on the overall structure of the carboprotein, as the altro-based carboprotein was found to be more alpha-helical than the corresponding galacto- and gluco-carboproteins. However, an influence on stability could not be detected in the present experiments, as the three carboproteins gave similar free energy of foldings (deltaG(F)H2O) and melting points in chemical and thermal denaturation experiments.     

An enantiomerically pure adamantylimido molybdenum alkylidene complex. An effective new catalyst for enantioselective olefin metathesis

An enantiomerically pure Mo-based complex that bears an alkylimido ligand is prepared and characterized through NMR spectroscopy and X-ray analysis. Mo complex 4 is the only reported chiral alkylimido catalyst; all previous chiral complexes are arylimido systems. These studies show that the chiral Mo catalyst exists exclusively as the syn isomer and that it offers unique reactivity and selectivity profiles in asymmetric olefin metathesis.     

A conjectural generalization of the n! result to arbitrary groups

The aim of this paper is to formulate a conjecture for an arbitrary simple Lie algebra g in terms of the geometry of principal nilpotent pairs. When g is specialized to sln, this conjecture readily implies the n! result and it is very likely that, in fact, it is equivalent to the n! result in this case. In addition, this conjecture can be thought of as generalizing an old result of Kostant. In another direction, we show that to prove the validity of the n! result for an arbitrary n and an arbitrary partition of n, it suffices to show its validity only for the staircase partitions.     

Transfer Matrices and Partition-Function Zeros for Antiferromagnetic Potts Models. II. Extended Results for Square-Lattice Chromatic Polynomial

We study the chromatic polynomials for m×n square-lattice strips, of width 9m13 (with periodic boundary conditions) and arbitrary length n (with free boundary conditions). We have used a transfer matrix approach that allowed us also to extract the limiting curves when n. In this limit we have also obtained the isolated limiting points for these square-lattice strips and checked some conjectures related to the Beraha numbers.     

Unusual Corrections to Scaling in the 3-State Potts Antiferromagnet on a Square Lattice

At zero temperature, the 3-state antiferromagnetic Potts model on a square lattice maps exactly onto a point of the 6-vertex model whose long-distance behavior is equivalent to that of a free scalar boson. We point out that at nonzero temperature there are two distinct types of excitation: vortices, which are relevant with renormalization-group eigenvalue 1/2 and non-vortex unsatisfied bonds, which are strictly marginal and serve only to renormalize the stiffness coefficient of the underlying free boson. Together these excitations lead to an unusual form for the corrections to scaling: for example, the correlation length diverges as J/kT according to Ae ² (1+be ^– +···), where b is a nonuniversal constant that may nevertheless be determined independently. A similar result holds for the staggered susceptibility. These results are shown to be consistent with the anomalous behavior found in the Monte Carlo simulations of Ferreira and Sokal.