310 



BIOCHEMICAL GENETICS 



heavy component might not be the same hemoglobin as that present in fresh 

 preparations. 



The physical structure of the hemoglobin molecule under various chemical 

 conditions has also been studied by ultracentrifugation. Field and O'Brien 358 showed 

 that human hemoglobin undergoes dissociation into half-molecules at pH 3.5-5.5, 

 and recently Hasserodt and Vinograd 524 described a similar phenomenon at pH 1 1 . 



Fig. 42. Comparison of murine oxyhemoglobins analyzed by 



ULTRACENTRIFUGATION. 



A B 



Patterns were photographed after 80 minutes of ultracentrifugation; phosphate buffer, 

 ionic strength 0.1, pH 7.5, 5° C. 



Left (A). Single hemoglobin of strain C57BL mice, S 20 value, ~4.7. 

 Right (B). Diffuse hemoglobin of strain 101 mice, S 2 o values. ~4.6 and 7.1. 



The dissociation is more complete at pH 1 1 , and less denaturation occurs. A dis- 

 advantage of dissociation at high pH, however, is that the heme-labelling methods 

 described by Singer and Itano 1210 cannot be used to establish transfer of hemoglobin 

 subunits during hybridization experiments, since methemoglobin of most species is 

 rapidly denatured by alkaline hydrolysis. However, Vinograd and Hutchinson 1345 

 have used C 14 -labeled hemoglobin to demonstrate molecular hybridization of 

 carbonmonoxyhemoglobins dissociated at pH 1 1 . 



