Wayne E. Steinmetz

A computer-assisted model for estimating protein secondary structure from circular dichroic spectra: Comparison of animal lactate dehydrogenases

Abstract A statistically valid method has been developed for estimating globular protein secondary structure from circular dichroic analysis. Circular dichroism data, in the form of mean residue ellipticity, at wavelengths between 210 and 240 nm provide an accurate and facile prediction of the fraction of helical structure in an uncharacterized globular protein. β-Strand can also be estimated indirectly using a different procedure once the fraction of helix has been determined. The analysis predicts a helical content of 42.1 ± 1.6, 42.7 ± 1.6, and 45.9 ± 1.6% for lobster tail and leg l -lactate dehydrogenase and Limulus skeletal muscle d -lactate dehydrogenase, respectively. Values of β-strand are calculated to be 28, 27, and 25% for the same enzymes. The results suggest a high degree of structural homology between the three enzymes and compare favorably to the X-ray-determined values of 41.6% helix and 25.8% β-strand for dogfish skeletal muscle lactate dehydrogenase.

On the Role of Polarizability in Chemical−Biological Interactions

This report considers the importance of electronic effects in their role in the QSAR of chemical-biological interactions. The problem of accounting for polarizability effects in ligand-substrate interactions is discussed in terms of molecular polarizability (MR) and NVE (number of valence electrons) using additive values for valence electrons. The two approaches give essentially the same result in examples of frog nerve toxicity and examples of nerve toxicity with rabbits and cockroaches. The point is made that no matter how one approaches QSAR, electronic interactions must be considered if we are to begin to develop a science of chemical-biological interactions.

Synthesis of a Molecular Charm Bracelet via Click Cyclization and Olefin Metathesis Clipping

We describe the synthesis of a polycatenated cyclic polymer, a structure that resembles a molecular charm bracelet. Ruthenium-catalyzed ring-opening metathesis polymerization of an amino-containing cyclic olefin monomer in the presence of a chain transfer agent generated an alpha,omega-diazide functionalized polyamine. Cyclization of the resulting linear polyamine using pseudo-high-dilution copper-catalyzed click cyclization produced a cyclic polymer in 19% yield. The click reaction was then further employed to remove linear contaminants from the cyclic polymer using azide- and alkyne-functionalized scavenging resins, and the purified cyclic polymer product was characterized by gel permeation chromatography, (1)H NMR spectroscopy, and IR spectroscopy. Polymer hydrogenation and conversion to the corresponding polyammonium species enabled coordination and interlocking of diolefin polyether fragments around the cyclic polymer backbone using ruthenium-catalyzed ring-closing olefin metathesis to afford a molecular charm bracelet structure. This charm bracelet complex was characterized by (1)H NMR spectroscopy, and the catenated nature of the small rings was confirmed using two-dimensional diffusion-ordered NMR spectroscopy.

NMR studies of the conformation of the natural sweetener rebaudioside A

Rebaudioside A is a natural sweetener from Stevia rebaudiana in which four beta-D-glucopyranose units are attached to the aglycone steviol. Its (1)H and (13)C NMR spectra in pyridine-d(5) were assigned using 1D and 2D methods. Constrained molecular dynamics of solvated rebaudioside using NMR constraints derived from ROESY cross peaks yielded the orientation of the beta-D-glucopyranose units. Hydrogen bonding was examined using the temperature coefficients of the hydroxyl chemical shifts, ROESY and long-range COSY spectra, and proton-proton coupling constants.

Preliminary spectroscopic characterization of six toxins from Latin American scorpions

This paper reports on spectroscopic studies of six toxins from the Latin American scorpions Centruroides noxius Hoffmann, Centruroides elegans Thorell and Tityus serrulatus Lutz and Mello. The isolation and purification of five of these toxins was described previously. The preparation of toxin II.9.2.2 from the venom of C. noxius is first described here. Circular dichroism and nuclear magnetic resonance spectra indicate similarities and differences between these scorpion toxins and previously characterized snake toxins. While there is evidence that the toxins from scorpions and snakes both contain extended beta-sheet secondary structures, the spectral properties of the scorpion toxins are overall of a different type from those of snake toxins. Among the six scorpion toxins those from T. serrulatus have spectral properties markedly different from those of the Centruroides species. Furthermore, thermal denaturation and amide proton exchange measurements showed that the globular structures of the Tityus toxins were markedly less stable and less rigid than those of the Centruroides toxins.

The conformational analysis of 1-haloalkanes by low resolution microwave spectroscopy

Abstract The low resolution microwave spectra of a series of 1-haloalkanes were studied. With the exception of 1-bromobutane only the all s-trans conformer and those gauche conformers in which a single dihedral angle is gauche were foud. By analyzing data from a homologous series, a gauche dihedral angle of 286 ± 4° was determined for the gauche conformers of the bromo and iodo species. An analysis of relative intensities of the 1-bromobutane and 1-iodobutane bands yielded standard Gibbs free energy differences.

Structure and Conformational Dynamics of Trichothecene Mycotoxins

A combination of NMR spectroscopy and molecular modeling has been employed to characterize the conformation and dynamics of the macrolide ring in verrucarin A and roridin A, two closely related toxins in the trichothecene mycotoxin family. Longitudinal carbon-13 relaxation times demonstrate the relative flexibility of the macrolide ring. The calculations, NOEs, and scalar vicinal coupling constants show that verrucarin A in CDCl 3 and CD 2Cl 2 predominantly adopts a single, well-defined conformation that matches the crystal structure. In contrast, roridin A is present as a mixture of two conformers.

3D QSAR study of the toxicity of trichothecene mycotoxins

Trichothecene mycotoxins, toxic natural products of fungi from the family Hypocreaceae, are potent inhibitors of protein synthesis. The application of 3D QSAR to these toxins explored the structural basis for their biological activities. A CoMFA (Q(2)=0.619, R(2)=0.921) model was developed for a set of 15 toxins with the trichothecene nucleus; CoMFA (Q(2)=0.518, R(2)=0.855) and CoMSIA (Q(2)=0.695, R(2)=0.960) models were developed for 31 toxins with the nucleus and a macrolide ring. The results show the role of electrostatics and steric factors in the activity of the toxins and indicate that the conformation of the macrolide ring influences the toxicity of the macrolide toxins.

Separation of electronic and hydrophobic effects for the papain hydrolysis of substituted N-benzoylglycine esters

The role of hydrophobic and electronic effects on the kinetic constants kcat and Km for the papain hydrolysis of a series of 22 substituted N-benzoylglycine pyridyl esters was investigated. The series studied comprises a wide variety of substituents on the N-benzoyl ring, with about a 300,000-fold range in their hydrophobicities, and 2.1-fold range in their electronic Hammet constants (sigma). It was found that the variation in the log kcat and log 1/Km constants could be explained by the following quantitative-structure activity relationships (QSAR): log 1/Km = 0.40 pi 4 + 4.40 and log 1/kcat = 0.45 sigma + 0.18. The substituent constant, pi 4, is the hydrophobic parameter for the 4-N-benzoyl substituents. QSAR analysis of two smaller sets of glycine phenyl and methyl esters produced similar results. A clear separation of the substituent effects indicates that in the case of these particular esters, acylation appears to be the rate limiting catalytic step.

NMR assignment and characterization of proton exchange of the ellagitannin granatin B.

Granatin B, a complex ellagitannin extracted from pomegranate fruit, has two equilibrating isomers, form a and form b. A full ensemble of proton and carbon‐13 NMR methods over a wide range of temperature enabled a complete assignment of the more abundant isomer and showed it to be form b. This result is based on the NMR data for granatin alone and agrees with the previous determinations which were based on a combination of chemical methods and a partial assignment of the NMR spectra. The new NMR spectra also yield exchange rates for the hydroxyl protons as a function of temperature. A molecular model involving hydrogen bonding provides an explanation for the exchange data. Copyright