30 



MICROSOMAL PARTICLES 



THE STABILIZATION OF RNA 



The RNA prepared in the manner described above exhibited values of sedi- 

 mentation constant and intrinsic viscosity that depended very much on the 

 solvent employed, and invariably these quantities slowly diminished with time. 

 Choosing 0.01 M phosphate buffer at pYL 7.0 as solvent, typical preparations 

 would have initially sedimentation constants of 9 and intrinsic viscosities of 0.6. 

 The sedimentation pattern as observed in ultraviolet optics was rather broad 

 and usually single peaked. The use of the above values in the Mandelkern- 

 Flory equation [6] gave approximately 200,000 as the molecular weight. 



In studying the gradual decay in molecular weight it was observed that heat- 

 ing to 60° C for a few minutes or the addition of KCNS to the extent of 0.2 M 

 would cause a 30 per cent fall in molecular weight. This indicates that the 

 polynucleotide chains of the original material were aggregated to a small but 

 significant extent. 



When the RNA was heated to progressively higher temperatures in the 

 0.01 M phosphate buffer the molecular weight was observed to continue falling 

 up to temperatures of about 80° C. After exposure to 80° the molecular weight 

 remained unchanged with time at room temperature, for prolonged periods at 

 83° and for short periods at 95°. Thus a single exposure to about 80° C pro- 

 duced a stabilization of the RNA. 



The changes which such heating induced in the sedimentation pattern are 

 shown in figure 2, where the sedimenting boundaries observed with ultraviolet 

 optics under identical conditions are shown for 0, 5, 15, and 45 minutes' heating 

 at 83° C. It is seen that the effect of the heating is to lower and to narrow the 



I oo 



75 



C /c 

 50 



0.25 



000 



6.0 



CHANGES IN SEDIMENTATION BOUNDARY 

 PRODUCED BY HEATING RNA TO 83°C 



Relotive concentration versus 

 distance from center of rototion 



Solvent -OIM KH 2 P0 4 - K 2 HP0 4 , pH 7 

 RNA concentration 38 if/cc. 



6.2 6 4 



DISTANCE I N CM. 



6 6 



Fig. 2. Effect of heating upon RNA sedimentation boundaries after 21 minutes at 

 59,780 rpm. 



