34. THE RIBONUCLEIC ACIDS OF VIRUSES 275 



Iii contrast to formaldehyde, Kethoxal does not cause a marked increase 

 in the UV absorption maximum of RNA or of mononucleotides. Only the 

 spectrum of guanylic acid is markedly changed after reaction with Kethoxal 

 and glyoxal. The product of the reaction of guanylic acid with the former 

 agent is very stable in contrast to the compounds resulting from the formal- 

 dehyde treatment of nucleotides. The spectrum of 1-methylguanine is not 

 altered after treatment with Kethoxal, and thus, the most likely place for a 

 reaction of the dicarbonyl compound is the 2-amino group and imino group 

 in position 1 of the guanine ring. When the reaction has gone to completion, 

 the amount of Kethoxal bound corresponds to less than half the number of 

 amino groups present in the RNA. This also speaks in favor of a synthetic 

 reaction with guanine groups. 



The reaction of infectious RNA with other chemical reagents. The reaction 

 of TMV-RNA with nitrous acid has been studied in great detail. 102 It leads 

 to well-defined reaction products. By measuring the rate of inactivation of 

 RNA by nitrous acid, it is possible to calculate the minimum number of 

 amino bases per mole of RNA necessary for the infectivity. Because of its 

 significance for the in vitro production of TMV mutants, this method will 

 be discussed in detail in the next chapter. Many other agents can react with 

 infectious RNA resulting in a loss of biological activity. The mode of action 

 of these agents is not yet known and the products of reaction have not yet 

 been identified. Several of these will be discussed, briefly, here. 



TMV or TMV-RNA can be rapidly inactivated by treatment with an excess of 

 dimethyl sulfate at neutral pH. 76 The reaction of dimethyl sulfate with virus results 

 in alterations of the protein which make this component insoluble so that it floccu- 

 lates. RNA is inactivated according to first-order kinetics if the methylation is per- 

 formed in a highly dilute alcoholic solution of dimethyl sulfate (molar ratio dimethyl 

 sulfate/RNA = 4000:1). Inactivation of RNA is probably due to a substitution of 

 the amino groups and the ring nitrogen atoms of the bases by methyl groups, since 

 the reaction products of the methylation of adenosine with dimethyl sulfate at 

 neutral pH are chiefly V 6 -methyladenosine and 1-methyladenosine. 103 



The bromination of RNA in aqueous solution, employing small amounts of bro- 

 mine, also leads to a rapid inactivatidn. 76 Sodium periodate, which is a specific 

 oxidizing agent of the czs-glycol group ( — CHOH — CHOH), also inactivates RNA if 

 the reaction is carried out at a weakly acid pH with an excess of NaI0 4 (20 moles 

 per mole RNA nucleotide). 76 



Hydroxylamine inactivates many RNA-containing animal viruses and also de- 

 stroys the infectivity of TMV-RNA at neutral pH. 104 



(4) Incorporation of " Unnatural" Bases in RNA. Certain purine and 

 pyrimidine compounds, which are structural analogs of the bases found in 



102 H. Schuster and G. Schramm, Z. Naturforsch. 13b, 697 (1958). 

 lu3 H. Bredereck, H. Haas, and A. Martini, Chem. Ber. 81, 307 (1948). 

 104 R. M. Franklin and H. Schuster, unpublished work (1959). 



