Shiro Matsuura

Electron microscopic studies on the biosynthesis of the 50 s ribosomal subunit in Escherichia coli

Two intermediary particles of the formation of 50 s ribosomal subunits, i.e. [30 s]- and 40 s particles, were isolated in 10−2 m-Tris-HCl containing 10−4 m-magnesium acetate at pH 7.8 and observed using an electron microscope. Most of the [30 s]-particles appear as elongated coils or thick thread-like molecules under the present experimental conditions, and in this respect they are similar to free ribosomal RNA. The 40 s particles resemble the 50 s ribosomal subunits, being much more spherical and compact than the [30 s]-particles. These observations suggest that the basic structure of the 50 s ribosomal subunit is formed mainly in the process from the [30 s]-particle to the 40 s particle.

Biosynthesis of cytochrome c and its posttranslational transfer into mitochondria.

Publisher Summary Cytochrome c is a peripheral membrane protein that is located in the intermembrane space of mitochondria, where it binds to cytochrome oxidase, an integral membrane protein. Cytochrome c represents an excellent model system for the study of the intracellular mechanisms by which proteins made on cytoplasmic ribosomes are sorted out and segregated to different compartments of subcellular organelles. In carrying out the posttranslational transfer experiments of newly synthesized cytochrome c, several precautions must be taken. Mitochondria should be very fresh and intact to prevent degradation of newly synthesized cytochrome c by proteinase released from mitochondria during the posttranslational incubation and to ensure its uptake by mitochondria. It is necessary to use proteinase inhibitors to protect newly synthesized polypeptides from proteolysis. However, the inhibitor effects on the posttranslational transfer of newly synthesized polypeptides should be examined. To rule out the possibility that newly synthesized cytochrome c binds to the cytoplasmic surface of outer mitochondrial membranes through ionic interaction, three experiments were performed—namely, effects of polylysine on the posttranslational transfer, resistance of newly synthesized cytochrome c to trypsin digestion, and conversion of apocytochrome c to holocytochrome c.