Escherichia wli ribosomes display a remarkable heterogeneity when submitted to polyacrylamide-gel electrophoresis (4O/, acrylamide). Both the 70-S ribosomes and 5 0 3 ribosomal subunits are resolved into a t least four and the 30-S ribosomal subunits into three particle subclasses.After reversal of the current, all particles refocus into one single band a t the origin. Ribosomes recovered from the gel after electrophoresis have retained some 40--100°/, of their capacity to synthesize peptide bonds. Their sedimentation behaviour has remained unaltered. Upon reelectrophoresis, no new peaks are detectable although the relative proportions of the various subclasses are frequently changed. The ratio in the occurrence of these subclasses is readily affected by a variety of factors, however, some of which are not directly related to the electrophoretic technique itself.The three classes of ribosomes and subunits (70-8, 5 0 3 and 30-S) display approximately equal electrophoretic mobilities when the sieving action is reduced to a minimum. This conclusion follows from experiments in which the ribosomes are submitted to electrophoresis in a sucrose gradient or in 0.4O/, agarose gels. I n contrast, electrophoresis in polyacrylamide gels with a gradient in pore size reveals the same ribosomal diversity as noted in gels with a fixed acrylamide concentration of 4O/,. I n these pore-gradient gels (3 -So/, acrylamide), the migration rate of each ribosomal subclass decreases continuously when the time of electrophoresis is prolonged. After about 30 h, further penetration seems to be blocked.All three classes consist of particles with low electrophoretic mobility (designated I particles) and those which move faster (designated I1 particles). The I particles are partially converted to I1 particles by incubation a t 37 "C and/or pelleting prior to elcctrophoresis. The I1 particles are converted to I particles upon raising the ribosome concentration. The electrophoretic profiles displayed by the various 50-S subclasses arc not strongly affected by variations in Mg2+ concentration in the range 0.5-10.0 mM. The electropherograms of 30-8 subunits become more diffuse when the Mg2+ concentration is raised. The possible occurrence of ribosomal dimers and/or conformation changes is discussed.Polyacrylamide-gel electrophoresis has proved to be a power-ful tool for fractionating various kinds of biologically important macromolecules like proteins [1,2], RNAs [3--51 and DNAs [el. Hjerten et al. [7] were the first to describe an electrophoretic separation of Escherichia coli ribosomal subunits in polyacrylamide gels. Later on, Dahlberg et al. [S] used composite gels of agarose and polyacrylamide to characterize bacterial polyribosornes, ribosomes and ribosomal subunits. Agarose can be omitted when only 70-S ribosomes and subunits are to be separated and we have used 40/, polyacrylamide gels to analyse ribosomal mixtures of E . coli after their interaction with the initiation factor IF-3 [9]. Dahlberg et al. [S] already noticed an unanti...
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