263 



Sakae Katoh and Atusi Takamiya 



GENERAL PROPERTIES OF PLASTOCYANIN 



Plastocyanin is readily extracted with dilute buffer solution from an 

 acetone powder of a leaf homogenate. It is purified by ammonium sulfate 

 fractionation and column chromatography with diethylaminoethyl cellulose^ >. 

 Crystallization of the protein from Chenopodium leaves has been achieved by 

 Yakushiji (personal communication). The molecular weight of the purified 

 protein was estimated from sedimentation and diffusion data to be 21, OOOw). 

 The protein is acidic in nature with an isoelectric point less than pH 4. 



The oxidized protein is deep blue in color and exhibits a complex absorp- 

 tion spectrum with three maxima at 460, 59 7 and 770 m^. The ultraviolet peak 

 shows vibrational fine structure bands due to the amino acid constituents. On 

 reduction, the blue color completely disappears and there is no absorption in 

 the visible and far red regions. The oxidized protein is readily reduced by 

 various reducing reagents such as ascorbaic acid, hydroquinone, sodium 

 hydrosulfite and reduced cytochrome c. On the other hand, the reduced form 

 of the protein is completely incapable of reacting with molecular oxygen through 

 oxidation or oxygenation. The oxidation reduction potential is 370 mv between 

 pH 5. 4 and 9.9. 



Plastocyanin contains about 200 amino acid residues and a small amount 

 of carbohydrate. The copper content was estimated to be 0. 58%, indicating 

 the presence of two atoms of copper. Titration with either PCMB or heavy 

 metal ions indicated that each copper atom is bound to the protein through the 

 sulfhydryl groups of cysteine. The original copper protein was reconstituted 

 by addition of an inorganic copper salt to the apoprotein prepared by acid 

 treatment. 



DISTRIBUTION OF PLASTOCYANIN IN PLANTS 



Plastocyanin was first observed in the aqueous extract of lyophylized 

 Chlorella elipsoidea (8). The green leaves of the following plants were found 

 to contain plastocyanin in significant amounts; spinach, parsley^ carrot, 

 turnip, crown daisy, Japanese scallion, Brassica campestris (Komatsuna), 

 Chenopodium album and Ulva sp.( 1°). Plastocyanin appears to occur ubiqui- 

 tously in the photosynthetic organisms. One exception was the photosynthetic 

 bacteria, since all attempts to detect a similar protein has failed. 



The occurrence of plastocyanin is limited to the green parts of the plants. 

 The protein is entirely absent from the white underground stem of Japanese 

 scallion and the roots of carrot and turnip, while the green leaves of these 

 plants contain significant amounts of the copper protein. This fact suggests a 

 close association of plastocyanin with the photosynthetic apparatus. In fact, 

 whole chloroplasts isolated from spinach leaves were found to contain plasto- 

 cyanin in a ratio of about 300 chlorophyll molecules per atom of copper of the 

 protein, or about 600 chlorophyll molecules per molecule of the protein. Since 

 the total copper content of spinach chloroplast was estimated to be approximately 



