Ribosomes from Gram-negative bacteria such as Escherichia coli exhibit non-specific translation of bacterial mRNAs. That is, they are able to translate mRNAs from a variety of sources in a manner independent of the "strength" of the Shine-Dalgarno region, in contrast to ribosomes from many Gram-positive bacteria, such as Bacillus subtilis, which show specific translation in only being able to t...

Eukaryotic ribosomes are much more labile as compared to their eubacterial and archael counterparts, thus posing a significant challenge to researchers. Particularly troublesome is the fact that lysis of cells releases a large number of proteases and nucleases which can degrade ribosomes. Thus, it is important to separate ribosomes from these enzymes as quickly as possible. Unfortunately, co...

The runoff of a mixture of "heavy" and "light" polysomes has been reported to yield hybrid ribosomes, of intermediate density. However, we found that the dissociation of free heavy ribosomes as they advance in a sucrose gradient causes a serious artefact in the demonstration of hybrid ribosomes. This artefact is eliminated by fixation with glutaraldehyde before sedimentation. Formation of hybri...

The intracellular ribosome crystals from chick embryos incubated at 5 degrees C have been isolated; stable products of their dissociation were shown to include single ribosomes, tetramers of ribosomes, and tetramers of large subunits. Conditions were found for the formation in vitro not only of the p4 crystals from ribosomal tetramers but also of the tetramers themselves from free ribosomes. Th...

Comparative studies of free ribosomal RNA and ribosomes were made with two probes, Mg++ ions and ethidium bromide, which interact with RNA in different ways. Mg++. E. coli 16 S rRNA and 30 S ribosomes were equilibrated with four different buffers. Equilibration required several days at 4 degrees and several hours at 37 degrees. In all buffers ribosomes bound more Mg than free rRNA, the differen...

In vitro studies of translation provide critical mechanistic details, yet purification of large amounts of highly active eukaryotic ribosomes remains a challenge for biochemists and structural biologists. Here, we present an optimized method for preparation of highly active yeast ribosomes that could easily be adapted for purification of ribosomes from other species. The use of a nitrogen mill ...