487 



M. B. Allen and J. C. Murchio 



gel. A typical membrane is shown in Figure 1. On formation of 

 membranes the absorption maximxun moves toward longer wavelengths, 

 from 672-673 m(j. to 677-678 mji. Previous studies ^3; indicate 

 that it is unlikely that this change is a scattering artifact 

 with our instnmentation. The membranes are astonishingly stable, 

 vigorous ultrasonic treatment being required for their 

 disintegration. 



The implications of this spontaneous membrane fonnation for 

 chloroplast structure remain to be explored. In the present 

 investigation, it raised the possibility that the photochemical 

 properties of the membranes might be different from those of the 

 presumed particle originally split from the chloroplast. We have 

 therefore been experimenting with techniques for purification and 

 concentration of P-672 that do not involve pelleting the material. 

 The most successful of these so far comprises passage through 

 a column of Sephadex G-200 after RNAse treatment, followed by 

 concentration with silica gel, stopping before the point of mem- 

 brane formation. This treatment removes most of the material 

 absorbing in the 260-280 m^ region, as well as whatever nat\iral 

 electron acceptors that react with the light induced free radical 

 are present in the crude P-672 preparation. The purified material 

 shows a distinct absorption peak at 3^0 m^i. The iraterial respon- 

 sible for this peak has not yet been identified. 



Electron micrographs of material thus purified^ prepared by the 

 spray droplet technique of Williams and Backus ^^^, are shown in 

 Figures 2, 3^ and 4. The preparations contain particles of var- 

 ious sizes, the smallest being 70-75 A, proceeding through 

 130-140 A, 200-210 A, to the largest, slightly greater than 300 A. 

 Small aggregates of these particles are also visible in the micro- 

 graphs. The particlf sizes observed suggest polymers of a fun- 

 damental unit of 70 A or less. However, control experiments have 

 shown that only about half of the particles in this preparation 

 contain chlorophyll, the remainder probably being ribosomal 

 contaminants. Until these can be removed it is not possible to 

 be assured that all the size classes represent chloroplast 

 particles. 



For the same reason, it is only possible to give order of 

 ma,gnitude figures for the chlorophyll content and molecular weight 

 of the particles. They contain 10-20^ chlorophyll and have an 

 average molec\ilar weight of the order of 1,000,000. The results 

 of papain treatment and extraction with fat solvents indicate a 

 lipoprotein complex, as might be expected. The possibility of the 

 presence of nucleic acids as an integral part of the complex is 

 being explored, but cannot, obviously, be settled until the 

 ribosomal contaminants are removed. 



