260 HEINZ SCHUSTER 



c. Size and Structure of the Infectious Ribonucleic Acid 



It was shown, in the previous chapters, that TMV has an RNA content 

 of 5 % which corresponds to a molecular weight of 2 X 10 6 for a particle 

 weight of 40 X 10 6 for the entire virus. An RNA molecule of this size must 

 consist of at least 6,000 mononucleotides of average molecular weight 320. 

 This value is obtained from the average molecular weight of all 4 nucleo- 

 tides (338) less one mole of water (18). 



The RNA obtained by phenol or detergent extraction of TMV is bio- 

 logically active. It is interesting to consider whether the entire RNA con- 

 tained in the TMV particle is necessary for the biological activity or whether 

 only part of this is needed. In other words, what is the minimum size of an 

 RNA molecule isolated from TMV which is still capable of initiating an 

 infection? Gierer 11, 68 studied this question by investigating the sedimenta- 

 tion and viscosity, as well as the kinetics of ribonuclease degradation, of 

 infectious RNA. He found that the total amount of RNA contained in a 

 virus particle is necessary for the biological activity. Therefore, RNA 

 molecules of a molecular weight smaller than 2 X 10 6 are biologically in- 

 active. 



(1) Determination of Molecular Weight from Sedimentation and Viscosity 

 Measurements. Stanley and Cohen 53 isolated a nucleic acid from TMV by 

 a short heat treatment in NaCl solution, and this RNA had a molecular 

 weight of approximately 300,000. It was, however, unstable and decomposed 

 further to units of molecular weight of approximately 60,000. Light-scat- 

 tering measurements on RNA, also prepared by heating a TMV solution, 

 gave a molecular weight of 1.7 X 10 6 under certain conditions of extrac- 

 tion. 69 



The RNA prepared from TMV by the phenol method is not homogeneous. 

 In the analytical ultracentrifuge, it is possible to observe a homogeneous 

 component of high molecular weight (component ^4) as well as an inhomo- 

 geneous component of smaller average molecular weight (component B). n ■ 68 

 It has not yet been possible to determine whether this smaller component 

 is also present in intact virus, or whether it arises from degraded virus 

 particles, or during the preparation of the RNA by degradation of larger 

 RNA molecules. The inhomogeneity of this component suggests that it 

 arises through degradation. The infectious RNA sediments have the same 

 velocity as component A, as could be demonstrated by testing supernatant 

 and sediment for infectivity after varying degrees of centrifugation. 



It is possible to determine the molecular weight from the sedimentation 

 constant and viscosity of a particular substance. Employing such data, the 



68 A. Gierer, Z. Naturforsch. 13b, 477 (1958). 



69 G. R. Hopkins and R. L. Sinsheimer, Biochim. et Biophys. Acta 17, 476 (1955). 



