374 MAHLON B. HOAGLAND 



ratio of about 1.8, is metabolically active as an. amino acid acceptor and as 

 an amino acid donor to protein under appropriate conditions. A very simi- 

 lar preparation may be obtained from whole yeast cells by direct treatment 

 with phenol. 126 Apparently the cell walls are permeable only to the relatively 

 smaller sRNA molecules since ribosomes are not extracted by this method. 

 Methods of isolation which involve very low pH's, or extraction with acidic 

 salts at 100°C. yield metabolically inactive sRNA. These methods may be 

 used, however, for assaying transfer RNA-amino acid compounds in reac- 

 tion mixtures since they do not cause separation of the amino acid from the 

 RNA. Preiss et al. m described a method for extracting transfer RNA from 

 E. coli cells by the use of hot detergent, followed by salt and alcohol frac- 

 tionation, and adsorption on and elution from charcoal. This product has 

 general characteristics similar to the material from other sources isolated 

 by the phenol method. 



(2) Molecular Weight and Base Composition. The only published sedi- 

 mentation constant for sRNA (from rat liver) is 1.85 Soo.w at a concentra- 

 tion of 0.003% in 0.15 M NaCl, 0.015 M citrate pH 6.8, 117 and this value, 

 with diffusion data, suggested a molecular weight of 15,000 to 20,000. Esti- 

 mates based on the number of chain ends of mammalian sRNA capable of 

 accepting amino acids or terminal nucleotides, and assuming no branching, 

 give molecular weight values of around 30,000. 117 ' 122 Direct analysis of 

 nucleosides released on alkaline hydrolysis also yields a similar result. 132 

 Somewhat higher values, of around 30,000 to 50,000, are estimated for the 

 E. coli transfer RNA preparation. 131 Yeast sRNA is reported to have a 

 sedimentation constant of 5, 126 which would be consistent with a molec- 

 ular weight closer to that of E. coli sRNA. 



Tissieres 125 has obtained a molecular weight of 27,000 for E. coli sRNA 

 based on sedimentation and diffusion measurements. The material was ob- 

 tained by phenol treatment of the upper two-thirds of a 3-hour, 78,000 g 

 supernatant fraction from an E. coli extract. The RNA gives a single elec- 

 trophoretic peak and the sedimentation constant is unaltered by versene 

 or salt. It is of considerable interest that the whole supernatant fraction 

 gave an identical sedimentation constant for sRNA (*S 20 ,„, = 4.0) as was 

 obtained with the phenol extracted material. This suggests that the sRNA 

 in the natural state is not bound to protein and exists free in the cytoplasm. 



Some preliminary data on the base composition of sRNA from vari- 

 ous tissues are given in Table HI. 133 - 136 Since methods of preparation 



131 J. Preiss, P. Berg, E. J. Ofengand, F. H. Bergmann, and M. Dieckmann, Proc. 

 Natl. Acad. Sci. U. S. 45, 319 (1959). 



132 D. B. Dunn, Biochim. et Biophys. Acta 34, 286 (1959). 



133 P. F. Spahr and A. Tissieres, J. Mot. Biol, 1, 237 (1959). 



134 J. F. Scott, unpublished data. 



