34. THE RIBONUCLEIC ACIDS OF VIRUSES 255 



neighboring pitches to each other in the RNA spiral. It is interesting that 

 the radial electron density distribution is similar in four strains of TMV 

 and in cucumber virus 4. 41 ' The strains differ from one another only in the 

 relative heights of some peaks in protein density, that is, in the weights of 

 protein lying at particular radii. 



The results obtained by X-ray analysis have been supported by other 

 studies. Schramm et a/. 46 were able to show that TMV is gradually degraded 

 at alkaline pH to protein and RNA. In this case, a 2 % TMV solution is 

 treated with 0.05 M sodium bicarbonate buffer at pH 10. Protein is gradu- 

 ally removed from the intact virus particle so that the naked RNA thread 

 can be observed in the electron microscope. 



Similar results were obtained by Hart 47 who treated TMV with the an- 

 ionic detergent, Duponol C, at an elevated temperature. In this case, the 

 protein seems to unravel from the end of the rod, since shortened rods may 

 be seen in the electron microscope. RNA fibers project from the end of 

 these shortened rods. Further heating of the virus in the presence of Dupo- 

 nol resulted in a shortening and finally in a disappearance of the rods ac- 

 companied by the appearance of naked fibers, many of which were 3000 A. 

 long. After treatment of the detergent-treated virus with ribonuclease, 

 only shortened rods, without any projecting fibers, could be found. This 

 demonstrated that the fibers were most likely the viral RNA. 



Schramm and Hart estimated the diameter of the RNA fiber visible in 



o 



the electron microscope to be 30-40 A., a value too small compared with 

 that obtained by X-ray crystallography. However, one must remember 

 that the drying of preparations for electron microscopic studies may lead 

 to an unavoidable shrinkage of the RNA fiber, especially since the virus 

 particle has a hollow core. It is possible to observe the core by treating 

 TMV with uranyl acetate 48 • 49 which diffuses into the core. The heavy metal 

 thus produces an electron dense core. 



The structural studies indicate that the viral RNA possesses a definite 

 "superstructure," namely a single helical chain resulting from its location 

 within the protein coat, which also protects the RNA from enzymic deg- 

 radation. 



In the following chapters, it will be shown how the RNA may be freed 

 from this nucleoprotein complex without loss of biological activity. Further- 

 more, the molecular structure of such RNA in solution will be discussed. 



45 K. C. Holmes and R. E. Franklin, Virology 6, 328 (1958). 



46 G. Schramm, G. Schumacher, and W. Zillig, Z. Natarforsch. 10b, 481 (1955). 



47 R. G. Hart, Viroloijg 1, 402 (1955); Proc. Natl. Acad. Sci. U.S. 41, 261 (1955). 



48 H. E. Huxley, Proc. Stockholm Conf. on Electron Microscopy, 1956 p. 260 (1957). 



49 H. Fernandez -Moran and G. Schramm, Z. Naturforsch. 13b, 66 (1958). 



