31 

 respectively) . Each protein eluted in the wash and was 

 applied to the fifth step of purification after samples were 

 removed for assay. 



DEAE KPO^ chromatography. DEAE columns were 

 equilibrated in 50 mM KPO4 buffer at pH 7.3, before applying 

 the proteins from the previous step. Gradients up to 0.3 M 

 KCl were applied to each column. Single peaks of each 

 protein were eluted, pooled and dialyzed into 50 mM Epps-KOH 

 containing 20% glycerol and 1 vnM DTT at pH 8.6. Samples 

 were saved, and the remaining proteins were subjected to the 

 sixth step of purification. 



2'5'ADP Sepharose-4B affinity chromatography. An 

 NADP*- specific 2'5'ADP Sepharose-4B affinity column was 

 prepared and equilibrated with the Epps-KOH buffer. The 

 SP-I sample was applied to the column and washed with 

 starting buffer before a 300-ml gradient from to 0.18 M 

 NaCl and to 12 @8,25 fiM NADP* was applied. The protein 

 eluted at about 4 fiM NADP* in the gradient. The SP-I 

 Activity peak was pooled and dialyzed into the Epps-KOH 

 buffer. A sample of SP-I was saved, and the remaining 

 fraction was used in the seventh step of purification. The 

 SP-II was applied to the affinity column similarly. 

 Purification of SP-II was completed with this step at a 

 final volume of 15 ml and total protein of 6 ng . 



