Demonstration and analysis of ribosomes and RNA associated with mitochondria of mammalian cells

Abstract

Mitochondrial fraction prepared from mammalian cells (rat liver) by high and slow speed centrifugation and purified by sedimentation in an isopycnic sucrose gradient are shown to contain ribosomes.These ribosomes have a sedimentation coefficient of 83 S (measured in the absence of KCl) and a buoyant density of 1.54 g/cm3 as compared to 80S and 1.55 g/cm3 for cytoplasmic ribosomes.The sedimentation behaviour of mitochondrial ribosomes varies as a function of KCl concentration. The dissociation of cytoplasmic and mitochondrial ribosomes into sub‐units begins towards 0.1 M KCl and is complete in 1 M KCl. The sedimentation coefficient of the intact ribosomes and their dissociated units fall as the concentration of KCl is increased between 0 and 1 M.RNA prepared from rat liver mitochondrial fraction contains a low molecular weight component and two high molecular weight species whose sedimentation coefficients 26 S and 17 S correspond to molecular masses of 1.51 × 106 and 0.61 × 106 daltons. These mitochondrial RNA can be separated by sucrose gradient sedimentation from cytoplasmic ribosomal RNA species sedimenting at 28 S and 18 S.Mitochondrial RNA were also characterized by acrylamide gel electrophoresis. In 2.50% gels, the electrophoretic mobility of high molecular weight mitochondrial RNA was found to be slightly higher than that of their cytoplasmic counterparts. Moreover, the RNA corresponding to the smaller units was separated into two distinct peaks.Electrophoresis in 8% acrylamide gels revealved 5 S and 4 S RNA with a migration pattern similar to that of cytoplasmic 5 S and transfer RNA but present in different amounts and additional RNA with an electrophoretic mobility of 0.31 and 0.85 if that of 4 S RNA is equalled to 1.