34. THE RIBONUCLEIC ACIDS OF VIRUSES 289 



g. Reconstitution of Virus from Ribonucleic Acid and Protein 



Before the experimental demonstration of the infectivity of TMV-RNA, 

 Fraenkel-Conrat and Williams, 136 and, later, Commoner et a/. 137 '• 138 de- 

 scribed the formation of infectious particles by mixing inactive protein, 

 which had been prepared by alkaline degradation of TMV at pH 10.5, 

 with nucleic acid which had been prepared by dodecyl sulfate. The two 

 components were inactive when tested separately on Nicotiana glutinosa. 



After mixing 1 % solutions of the two components in the proportion of 10 

 parts protein to 1 part RXA and incubating at 3°C. for 24 hours (pH 6.0- 

 7.0), nucleoprotein particles carrying virus activity were found. The nucleo- 

 protein particles appeared to be identical in shape and size with TMV ex- 

 cept for a greater randomness in length. They contained 5-6 % RNA as do 

 TMV particles and were not attacked by ribonuclease. Upon treatment 

 with detergent, there was a partial degradation of virus protein exactly as 

 in the case of native TMV. A central RNA strand was visible after this 

 treatment. The particles proved somewhat more labile to detergent than 

 TMV. 



The combination of protein and nucleic acid fractions isolated from dif- 

 ferent strains of TMV (the so-called mixed reconstitution) was also pos- 

 sible. 139, 140 RXA from 4 different strains of TMV was recombined with the 

 protein component from 3 of these strains. All combinations of recombined 

 nucleoprotein were biologically active. In every case, the reaction products 

 gave the same disease symptoms as did the strain supplying the RXA. The 

 serological characteristics of mixed virus preparations were shown to re- 

 semble those of the protein component. 



After the preparation of infectious RNA, it became clear that the forma- 

 tion of a biologically active virus could not have resulted from a combina- 

 tion of inactive components. Instead, the comparatively low activity of the 

 RNA component is, simply, significantly increased by addition of the virus 

 protein. This interpretation is also supported by the fact that the activity 

 of TMV particles, which had been partially degraded with detergents, can 

 be increased by about tenfold after addition of native virus protein. 141 The 

 biological activity of the detergent degraded particles could be further 

 reduced by ribonuclease, whereas the particles which had been incubated 

 with protein were resistant to ribonuclease. 



136 H. Fraenkel-Conrat and R. C. Williams, Proc. Natl. Acad. Sci. U.S. 41, 690 

 (1955). 



137 J. A. Lippincott and B. Commoner, Biochim. et Biophys. Acta 19, 198 (1956). 



138 B. Commoner, J. A Lippincott, G. B. Shearer. E. E. Richman, and J. H. Wu, 

 Nature 178, 767 (1956). 



139 H. Fraenkel-Conrat, J. Am. ('hem. Soc. 78, 882 (1956). 



140 H. Fraenkel-Conrat and B. Singer, Biochim. et Biophys. Acta 24, 540 (1957). 



141 R. G. Hart, Nature 177, 130 (1956). 



