Two-dimensional crystalline sheets of Bacillus stearothermophilus 50S ribosomal subunits containing a nascent polypeptide chain

Polylysine chains were synthesized on Bacillus stearothermophilus ribosomes in a poly(A)-programmed in vitro system. After separation of the ribosomal subunits by sucrose gradient centrifugation, the polylysine chains (in contrast to the polyphenylalanine chains synthesized in a poly(U) system) reproducibly remained attached to the large ribosomal subunit. It was possible to produce two-dimensional crystalline sheets from the large ribosomal subunits containing the polylysine chains. These sheets are an essential prerequisite for three-dimensional reconstruction studies aiming to show that the tunnel in the large ribosomal subunit provides a path for the nascent polypeptide chain

Three-dimensional image reconstruction from ordered arrays of 70S ribosomes

A better understanding of the molecular mechanism of protein biosynthesis still awaits a reliable model for the ribosomal particle. We describe here the application of a diffraction technique, namely three-dimensional image reconstruction from two-dimensional sheets of 70S ribosomes from Bacillus stearothermophilus at 47 Å resolution. The three-dimensional model obtained by these studies shows clearly the two subunits, the contact points between them, an empty space large enough to accomodate the components of protein biosynthesis, the location of regions rich in RNA and a possible binding site for mRNA. The tunnel within the 50S particle which may provide the path taken by the nascent polypeptide chain in partially resolve

The growth of ordered two-dimensional sheets of 70 S ribosomes from Bacillus stearothermophilus

Well ordered two-dimensional sheets of intact 70 S ribosomes from Bacillus stearothermophilus have been obtained in vitro using salt-alcohol mixtures. These sheets consist of relatively small unit cells with dimensions of 200 + 20 A and 400 f 30 A. Diffraction patterns of electron micrographs of these sheets stained with uranyl acetate contain features to 42 A resolution. Ribosome Two-dimensional sheet 70 S ribosomal particle (Bacillus stearothermophilus

The growth of ordered two-dimensional sheets of 70 S ribosomes from Bacillus stearothermophilus

Well ordered two-dimensional sheets of intact 70 S ribosomes from Bacillus stearothermophilus have been obtained in vitro using salt-alcohol mixtures. These sheets consist of relatively small unit cells with dimensions of 200 + 20 A and 400 f 30 A. Diffraction patterns of electron micrographs of these sheets stained with uranyl acetate contain features to 42 A resolution. Ribosome Two-dimensional sheet 70 S ribosomal particle (Bacillus stearothermophilus

Characterization of crystals of small ribosomal subunits

Crystals of intact small ribosomal subunits from Thermus thermophilus have been obtained from functionally active particles. The crystals (P42$_1$2, 407 Å × 407 Å × 171 Å) are suitable for X-ray crystallography analysis to 9·9 Å using synchrotron radiation at cryotemperature. Crystallographic data from native and a potential heavy-atom derivative have been collected

Role of cGMP Mechanisms in Response of Rat Pulmonary Arteries to Hypoxia

We have demonstrated previously that in response to hypoxia, isolated rat pulmonary arteries show an initial endothelium-dependent relaxation followed by an endothelium-independent transient contraction. In the presence of increased extracellular Ca2+, both of these responses were enhanced in endothelium-intact arteries. Nitro-L-arginine, a blocker of the biosynthesis of endothelium-derived relaxing factor (EDRF), abolished the initial endothelium-dependent relaxation and Ca(2+)-induced enhancement of hypoxic contraction in endothelium-intact arteries but did not alter responses in endothelium-denuded vessels. Inhibition of prostaglandin formation with indomethacin had no effect on the hypoxia-elicited responses. Preincubation with LY 83583, an inhibitor of guanylate cyclase activation, abolished the initial hypoxia-elicited relaxation and subsequent contraction. M & B 22948, a guanosine 3\u27,5\u27-cyclic monophosphate (cGMP) phosphodiesterase inhibitor, decreased tone under O2 but not under N2, causing an apparent enhancement of the contraction to hypoxia. Thus the modulation of hypoxic responses by the endothelium is dependent on changes in EDRF production, and a decrease in smooth muscle cGMP not involving an EDRF mechanism appears to mediate the endothelium-independent hypoxic contraction observed in the isolated rat pulmonary artery