274 



E. P. Katz, G. Mechanic, M. J. Glimcher 



peaks are strongly dependent on protein concentration, solvent, pH and the ionic 

 strength of the solution. In so short a paper all these variables on the various para- 

 meters characterizing the system are impossible to present or discuss so only repre- 

 sentative data will be presented here. Typical sedimentation velocity experiments at 

 various protein concentrations at pH 7 .7 , 0.1 M NH4HCO3 is shown in Fig. 1. Three 

 peaks are evident in Fig. 1 a (1.4''/o protein) and the apparent distribution of the 

 components is such that 80"/o is represented by the fast moving peak and about IC/o 

 by each of the slower moving peaks. The Son -w value of the major peak is about 9 S, 

 while the two minor peaks have Sooav values of approximately 2 S and 1 S respec- 

 tively (Fig. 1 e). 



As the total protein concentration is decreased there is a redistribution of the 

 material so that progressively less of the protein is represented by the faster sediment- 

 ing peak. This is demonstrated in Fig. 1 b, c and d where we see sedimentation 

 patterns of O./o/o, 0.350/o and ClZSVo protein solutions at pH 7 .7 in 0.1 M 

 NH4HCO3 . Over the concentrations presented here, not only does the apparent 

 ratio of fast to slow moving species change by a factor of 2.4, but also the calculated 

 area of the composite 1 — 2 S peaks changes with total protein concentration. In other 

 experiments it was found that the relative proportions of the two slower moving 

 peaks appear to be relatively insensitive to total protein 

 concentration. 



The schleiren patterns in Fig. 1 also demonstrate that 

 the fast sedimenting peak is not resolved from the slower 

 moving ones. This indiates that the three peaks seen 

 apparently constitute one continuous boundary. 



At pH's 8.82, 8.3 and 6.04 the sedimentation velocity 

 patterns are similar to those seen in Fig. 1. At pH's 6.04, 

 however, there is a progressive redistribution as more of 

 the material appears in the slower sedimenting peaks. 

 The Soo.w values of the fast moving peaks changes ra- 

 dically with pH and ionic strength as seen in Fig. 2. 



It may also be noted here that high concentrations 

 of urea deaggregate the protein and only slow moving 

 boundary of approximately 1 S may be seen in sedi- 

 mentation velocity patterns. Upon dialysis against neu- 

 tral buffer the patterns revert back to their original 

 form shown in Fig. 1 a. 

 The electrophoretic patterns of a 1.4"/o protein solution at various pH's are 

 shown in Figs. 3 and 4. 



Lowering the protein concentration does not change the distribution of the 

 proteins in either the ascending or descending pattern. At pH 6.04 (Fig. 3 c) the 

 major boundary remains with the ^ and f boundaries indicating the most of the 

 protein is in an isoelectric condition. 



At pH 3.61 (Fig. 4 f) there is a marked dift'erence in the patterns obtained from 

 the two arms of the electrophoretic cell. The pattern in which protein is migrating 

 into solution shows a number of hypersharp peaks (ascending pattern) whereas the 

 material migrating through protein solution gives diffuse boundaries (descending 

 pattern). 



Fig. 2. Plot of SSo.w at V 

 pH's at 0.1 M and 0.25 M 



8 pW 9 

 rlous 



