Biophysical studies on circle formation by Sindbis virus 49 S RNA

Abstract

Sindbis virus 49 S RNA extracted from virions is completely in the form of hydrogen-bonded circles, which are separable from linear 49 S RNA in sucrose gradients containing 0·01 M-NaCl. The melting temperature (t_m) of the circles is 39·5°C in 0·02:3 M-NaCl and 53·5°C in O· l M-NaCl. The ΔH value for cyclization is - 160 kcal/mol and the ΔS approximately -500 entropy units. These parameters indicate that the length of the double-stranded region which is formed upon cyclization of the molecule is most likely short, in the order of 10 to 20 nucleotides. Our data indicate that extensive mismatching in this double-stranded region is unlikely. Intact linear 49 S RNA molecules readily renature to form circles under appropriate conditions, the energy of activation for this process being 42·6 kcal/mol. From the measured rate constants for cyclization, it is clear that Sindbis RNA will form circles readily under physiological conditions of temperature and ionic strength. The virion RNA from Semliki Forest virus also forms circles whose t_m value is very similar to that of Sindbis RNA circles, suggesting that the sequences involved in cyclization have been conserved.

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