Solid-state 15N NMR of oriented lipid bilayer bound gramicidin A'

Highly oriented samples of lipid and gramicidin A' (8:1 molar ratio) have been prepared with the samples extensively hydrated (approximately 70% water v/w). These preparations have been shown to be completely in a bilayer phase with a transition temperature of 28 degrees C, and evidence is presented indicating that the gramicidin is in the channel conformation. An estimate of the disorder in the alignment of the bilayers parallel with the glass plates used to align the bilayers can be made from the asymmetry of the nuclear magnetic resonances (NMR). Such an analysis indicates a maximal range of disorder of +/- 3 degrees. Uniformly 15N-labeled gramicidin has been biosynthesized by Bacillus brevis grown in a media containing 15N-labeled Escherichia coli cells as the only nitrogen source. When prepared with labeled gramicidin, the oriented samples result in high-resolution 15N NMR spectra showing 12 resonances for the 20 nitrogen sites of the polypeptide. The frequency of the three major multiple resonance peaks has been interpreted to yield the approximate orientation of the N-H bonds in the peptide linkages with respect to the magnetic field. These bond orientations are only partially consistent with the extant structural models of gramicidin.     

The renin-angiotensin system in hypothyroidism of short duration

In six hypothyroid patients (2 male, 4 females, ages 22 through 59 years), plasma renin activity (PRA) and aldosterone (Aldo) were measured when the patients were euthyroid on levothyroxine therapy and one month after the therapy was stopped. Colonic mucosal potential differences were measured during the hypothyroid and euthyroid stages, and catecholamine sensitivity was determined by the blood pressure response to infused norepinephrine. Significant differences were observed in the PRA and aldosterone concentrations which were 4.1 +/- 2.5 ng/ml/h and 9.4 +/- 5.9 ng/dl, respectively in the hypothyroid stage and 6.9 +/- 2.3 ng/ml/h and 15.2 +/- 7.3 ng/dl, respectively when the patients were made euthyroid. The colonic mucosal potential differences (which reflect increased endogenous mineralocorticoid activity), became more electronegative after correction of hypothyroidism (-16.8 +/- 7.5 mV vs -32 +/- 18.2 mV; P less than 0.04) concentrations. Statistically significant decreases in norepinephrine pressor effects were observed in hypothyroid patients when compared to the euthyroid state (7.4 +/- 2.3 vs 10.9 +/- 1.9 micrograms/ng/min; P less than 0.01). It is concluded that patients with hypothyroidism have a hormonal pattern reminiscent of "low renin hypertension", and exhibit decreased sensitivity to catecholamines. Such changes are corrected when the patients become euthyroid on levothyroxine therapy.     

Sequences of E/NS1 Gene Junction from Four Dengue-2 Viruses of Northeastern Thailand and Their Evolutionary Relationships with Other Dengue-2 Viruses

We determined the 240-nueleotide sequences of the E/NS1 gene junction of four dengue-2 viruses by the primer extension dideoxy chain termination method. These viruses were isolated from dengue patients with different clinical severities in Nakhon Phanom, Northeastern Thailand in 1993. The results were compared with the 52 published dengue-2 sequences of the same gene region. Sequence divergence of four new isolates varied from 4.17% to 5.42% compared with dengue-2 prototype New Guinea C strain whereas it varied from 5.42% to 6.67% and from 6.67% to 7.09% when compared with Jamaica 1409 strain and PR159/S1 strain, respectively. All nucleotide substitutions were found at the 3rd position of the codons which were silent mutations. All 56 isolates studied were classified into five genotypic groups by constructing the dendrogram. The results indicated that four new isolates from Northeastern Thailand belong to genotype II of dengue virus serotype 2, and were most closely related to prototype New Guinea C strain. We also observed the variation in nucleotide and amino acid sequences among clusters of isolates (Thailand-1980, Malaysia-1989 and Thailand-1993) which were obtained from the dengue patients with different clinical severities. The significance of these genetic differences have been discussed in terms of the possible correlation between genetic variability and virulence.     

3-Azi-1-methoxybutyl D-maltooligosaccharides specifically bind to the maltose/maltooligosaccharide-binding protein of Escherichia coli and can be used as photoaffinity labels

Maltooligosaccharides with two to six (alpha 1-4)-linked glucose residues, carrying at their reducing end a 3-azi-1-methoxybutyl group in either alpha or in beta glycosidic linkage, were synthesized. These maltooligosaccharide analogues inhibit maltose uptake via the maltose-binding-protein-dependent transport system in Escherichia coli. The concentration of half-maximal inhibition of maltose transport, at 15 nM concentration, decreases with increasing chain length of the analogue, levelling off at 40 microM after a chain length of four glucose residues in the alpha series and at 350 microM after a chain length of three glucose residues in the beta series. The inhibition of maltose transport occurs at the level of the periplasmic maltose-binding protein. 3-Azi-1-methoxybutyl alpha-D-[3H]maltotrioside was bound by the maltose-binding protein with a Kd of 0.18 mM. Irradiation at 350 nm of purified maltose-binding protein in the presence of 4 microM of this substrate labeled the protein covalently; labeling was prevented by 1 mM maltose. Using a crude preparation of periplasmic proteins two proteins were labeled, the maltose-binding protein and alpha-amylase. Thus, 3-azi-1-methoxybutyl alpha-D-maltooligosaccharides are potent photoaffinity labels for proteins with maltooligosaccharides-binding sites.     

Evolutionary origins of taro (Colocasia esculenta) in Southeast Asia

As an ancient clonal root and leaf crop, taro (Colocasia esculenta, Araceae) is highly polymorphic with uncertain genetic and geographic origins. We explored chloroplast DNA diversity in cultivated and wild taros, and closely related wild taxa, and found cultivated taro to be polyphyletic, with tropical and temperate clades that appear to originate in Southeast Asia sensu lato. A third clade was found exclusively in wild populations from Southeast Asia to Australia and Papua New Guinea. Our findings do not support the hypothesis of taro domestication in Papua New Guinea, despite archaeological evidence for early use or cultivation there, and the presence of apparently natural wild populations in the region (Australia and Papua New Guinea).