THE STRUCTURE OF VIRUSES 



125 



phosphate buffer at pH 7 on tobacco-mosaic 

 virus has been studied in detail, and the 

 virus was found to be rapidly disinte- 

 grated into low-molecular-weight protein 

 components free of nucleic acid, with loss 

 of virus activity, serological specificity, and 

 ability to show stream double refraction. 

 The denaturation was accompanied by the 

 appearance of sulfhydryl groups, as indi- 

 cated by a positive nitroprusside reaction, 

 and the low-molecular-weight protein which 

 was formed was soluble in water but in- 

 soluble in 0.1 M or more concentrated salt 

 solutions. 



The fact that the virus preparations that 

 have been isolated have consisted essenti- 

 ally of high-molecular-weight protein ma- 

 terial has been utilized in still another type 

 of experimental approach. The materials 

 can be sedimented from solution by means 

 of a high-speed centrifuge because of their 

 unusually high molecular weight, and, since 

 they are protein and have an isoelectric 

 point, the sedimentation of negatively 

 charged protein, of positively charged pro- 

 tein, and of neutral or uncharged protein 

 can be studied. In the case of tobacco- 

 mosaic virus, it was found that regardless 

 of the charge the protein and the agent 

 carrying the virus activity sedimented at 

 exactly the same rate. This is good evi- 

 dence that the activity is a property of the 

 protein, for, if the activity were due to a 

 small amount of some material of a differ- 

 ent size mixed with the protein, it is obvi- 

 ous that under one of the conditions cited 

 the mixture would have been separated and 

 the activity would not have been found 

 with the high-molecular-weight protein. 

 By the high-speed centrifugation of mix- 

 tures of tobacco-mosaic virus with egg 

 albumin, globin, trypsin, or pepsin, it w^as 

 found possible to separate the character- 

 istic high-molecular-weight nucleoprotein 

 with unchanged activity. Gratia and Manil 

 obtained similar results with mixtures of 

 tobacco-mosaic virus and a bacteriophage. 



Neurath and Saum, using the refracto- 

 metric method of Lamm, found the diffu- 

 sion constant of chemically purified to- 

 bacco-mosaic virus to be about 3x10"* 



square cm per second, and Frampton, using 

 the same method, obtained a value of 2.1 x 

 10~^ square cm per second. The diffusion 

 constant obtained by Hills and Vinson by 

 means of the Northrop-Anson diffusion cell 

 is probably far too large, since these work- 

 ers did not use sufficient electrolyte to eli- 

 minate the accelerating effect of small ions 

 on the virus. In most of their experiments, 

 virus was permitted to diffuse from a dilute 

 electrolyte solution into distilled water, and 

 in the remaining experiments, from a 

 trypsin solution into a trypsin-free solu- 

 tion. Purified preparations of tobacco- 

 mosaic virus have been subjected to a care- 

 ful immunological study; when sufficiently 

 purified it was not found possible to detect 

 material other than the virus protein even 

 by the sensitive precipitin and anaphylactic 

 tests. Evidence of a different nature but 

 also indicative of homogeneity was provided 

 by the results obtained by means of the 

 analytical ultracentrifuge and the electro- 

 phoresis apparatus, for in each case a single 

 sharp boundary characteristic of a single 

 molecular species was obtained. It was 

 found impossible to separate virus activity, 

 from protein by filtration through collodion 

 or other types of filters. The ultraviolet- 

 light-absorption spectrum of purified to- 

 bacco-mosaic virus preparations was found 

 to agree essentially with the destruction 

 spectrum of virus activity, thus indicating 

 a close relationship between the two. 



A great amount of experimental work on 

 the purified virus preparations has already 

 been completed, and an even larger volume 

 is now in progress. Still more viruses are 

 being obtained in purified form from time 

 to time and ever-increasing amounts of the 

 ones already purified are being made avail- 

 able for experimentation. In all of the 

 work that has been reported to the present 

 time, or that is known to me, not a single 

 bit of experimental evidence has been ob- 

 tained that is incompatible with the idea 

 that the various purified and unaltered 

 virus preparations, or at least their chief 

 components, actually consist of the active 

 agent. On the contrary, there is an im- 

 posing array of evidence which indicates 



