12 



MICROSOMAL PARTICLES 



gation. Potassium phosphate buffer, pH 7.0 and ionic strength 0.1, was used in 

 the preparation of extracts. 



Preparative and analytical sedimentation were carried out in Spinco ultra- 

 centrifuges Models L and E, respectively. A diffusion constant measurement 

 was made by free diffusion in a Claesson cell [4]. Partial specific volume was 

 determined by density measurements in a Lipkin pycnometer [5]. 



RESULTS 



Decomposition Inhibitor in E. coli Extracts. It has been shown that the ex- 

 tractive procedure described here yields solutions that are highly reproducible 

 as determined by sedimentation and electrophoretic behavior [6]. The sedi- 

 mentation diagram (see fig. 1) corresponds closely to those obtained previ- 

 ously [2] by other methods of cell disruption. The rapidly sedimenting 40 S 

 component is clearly resolvable, and it appears that a considerably purified prep- 

 aration should be obtainable simply by successive differential centrifugation. 

 In early fractionation attempts, pellets from solutions subjected to centrifugal 

 fields about 100,000^ for 90 minutes (in the no. 30 rotor of the Model L Spinco) 

 were found to be only partly resoluble, in agreement with the finding of 

 Schachman et al. [2]. Moreover, the soluble material, as shown in figure 1, 

 contained an unexpectedly large amount of more slowly sedimenting material 

 along with a disappointingly low quantity of 40 S component. 



In a subsequent study to determine the effect of dialysis on the nature of 

 E. coli extracts, an observation was made that proved to be an important step 

 in this purification problem. As shown in figure 2, the 40 S component in 

 dialyzed extracts was greatly reduced in concentration with a simultaneous in- 

 crease in the quantity of more slowly sedimenting material. In additional ex- 

 periments, however, the effect was found to be diminished as the dialysate- 

 extract volume ratio was decreased. This suggested the presence, in those 

 extracts, of a dialyzable material that functions as a stabilizer of the 40 S com- 

 ponent. A substance with an analogous property has been reported by Peter- 

 mann and Hamilton [7] in studies with rat liver homogenates. 



Extract 



Pellet fraction 



Fig. 1. Sedimentation diagrams of an E. 

 coli extract and the pellet fraction derived 

 by a two-step centrifugation at 100,000^ for 

 90 minutes. Patterns were recorded 7 min- 

 utes after attainment of full field strength, 

 about 250,00%. 



Extract 



Extract dialyzed 

 against buffer 



Fig. 2. The effect of dialysis on the sedi- 

 mentation behavior of an extract from E. 

 coli. Recording of diagrams took place 8 

 minutes after full field strength was reached, 

 250,000^. 



