The kinetics of renaturation of denatured 5-S RNA from Escherichia coli have been studied by estimating the composition of reaction mixtures from absorbance profiles of stained zones containing the two forms separated by polyacrylamide-gel electrophoresis. The kinetics are first order and proceed to completion irrespective of temperature or Mga+ ion concentration, but they depend strongly on temperature. Linear Arrhenius plots yield an activation energy of 65 kcal/mol RNA. It is suggested that about nine base pairs of the denatured form dissociate and about the same number reform in a different configuration in the renatured conformer. Aubert et al. [5] have shown that 5-S RNA can exist in a t least two forms. These may be separated by chromatography on Sephadex G-100 or methylated bovine-serumalbumin-coated Kieselguhr [5] or electrophoretically on polyacrylamide gel [6] (and unpublished results). The form released from the 50-5 ribosomal subunit can participate in reconstitution, is termed the native form and can be partially converted to a denatured form by treatment with urea and EDTA [5]. The denatured form is chromatographically and electrophoretically distinct from the native form and has different spectroscopic properties [5--81. It is not a single conformational species since it may be resolved by polyacrylamide-gel electrophoresis into two components (unpublished results). On treatment with Mgz+ ions a t 60°C the denatured form is converted to a renatured form which is chromatographically and electrophoretically indistinguishable from the native form; it has closely similar optical properties [S] and can participate in the reconstitution of 50-8 ribosomal subunits [5]. Nevertheless it is claimed that the native and renatured forms are not identical in that the activity of the renatured form in reconstitution experiments is less than that of the native form [5] and that small differences in their optical properties ace discernible [S].
Jordan [9]has investigated the secondary structure of the native and denatured forms by partial digestion with nucleases and shown that the patterns of oligonucleotides produced are very different. It was concluded that the points of initial attack by the enzymes were not the same and inferred that the two forms had different secondary structures or base-pairing schemes. These observations do not preclude the possibility that they also differ in their tertiary structure. The renatured form has not yet been studied in this manner but in so far as the native and renatured forms are closely similar in many of their properties it is very likely that the denatured and renatured forms also differ in their secondary, and perhaps tertiary, structure.We have exploited, for the purposes of this investigation, the ability of polyacrylamide-gel electrophoresis to resolve the denatured and renatured [6] (and unpublished results) forms in order to study the effect of temperature and Mg2+ ion concentration on the rate of renaturation. This technique is particularly useful in this con...
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