STRUCTURAL BASIS OF RIBONUCLEASE ACTIVITY 



131 



SEDIMENTATION STUDIES ON RIBONUCLEASE 

 AND OXIDIZED RIBONUCLEASE 



O 

 X 



2.2 



2.0 



</) 



o 



1.0 



• NATIVE 

 oOXIDIZED 



1 



1.0 2.0 30 



G/IOOcc. 



Fig. 2. Sedimentation studies on ribonuclease and cxidized ribonuclease. 



ribonuclease, the oxidized molecule being much more dependent on concentra- 

 tion changes. 



Careful viscosity studies have also been carried out by Harrington and Schell- 

 man (1956), at the Carlsberg Laboratories, on native and oxidized ribonuclease, 

 both in 0.1 ionic strength buffer and in 8 ]M urea. Fig. 3 summarizes these vis- 

 cosity studies. Of particular interest here is the fact that the intrinsic viscosity 

 of native ribonuclease is greatly increased in 8 M urea, going from 3.3 to almost 

 9, and that the intrinsic viscosity of oxidized ribonuclease, which is now pre- 

 sumably completely free of covalent cross-linkages, is not a great deal larger, 

 being about 11.5. These studies suggest that the folding of the native ribonu- 

 clease chain, although restricted by four disulphide bridges, is arranged in such 

 a way that rupture of hydrogen bonds can lead to extensive disorientation, 

 nearly approaching that of the oxidized chain which presumably exists as a 

 random coil in solution. Among the hydrogen-bonded linkages which must be 

 present in native ribonuclease, one may certainly list those of the type described 

 by Pauling, Corey and Branson (1951), and perhaps further hydrogen 

 bonds of the sort originally suggested by Crammer and Xeuberger (1943) in- 

 volving linkages between hydroxyl groups of tyrosine side-chains and free 



