657 



N. E. Tolbert 



unpublished). Below 207o oxygen, glycolate synthesis ceases. 

 Thus the conditions for glycolate synthesis are those which occur 

 in nature — high O2 and low COg . Glycolate oxidase, a FMN con- 

 taining enzyme, in vitro has a low affinity for oxygen, so high 

 oxygen concentration would lead to the oxidation of glycolate 

 rather than accumulation. Thus aerobic conditions favor both gly- 

 colate synthesis, and because of the oxidase, its metabolism. 

 Glycolate oxidase can also be coupled to DCPIP or to a naturally 

 occurring quinone-like material which we have not identified. 

 The nature of the oxygen requirement for glycolate production is 

 unknown. Warburg has suggested that aerobic oxidation of a C2 

 product of photosynthesis to glycolate must occur first in order 

 that O2 evolution can generate reducing power (5). 



Effect of phosphate : The impairment of photosynthetic activ- 

 ity by washing algae with water is well recognized. Three wash- 

 ings of Chlamydomonas reinhardti reduced their photosynthetic 

 ability 80% as measured by C^'^Oa fixation (Table 4). This inhi- 

 bition was completely restored by 3 x 10"^ M phosphate, phospho- 

 glycolate or phosphoglycerate (39). Glucose-6-P was much less 

 effective and ATP was ineffective. However, certain other non- 

 phosphate compounds such as serine also restored the photosynthe- 

 tic activity, which is indicative of the complexity of the phe- 

 nomenon. 



Table 4. Effect of washing and phosphate on COg 

 fixation by Chlamydomonas. 



Total c/s in 10 min. by 1 ml aliquot 



Additions '^ Unwashed algae Washed algae 



none 2,700 546 



3.3 X 10"3M phosphate 2,960 3,100 



3.3 X 10"^M phosphoglycolate 2,560 3,050 



■"-All solutions were adjusted to pH 7. 



The products of C^^02 fixation depend on the phosphate in the 

 medium. Very fast labeling of glycolate occurs in algae when 

 photosynthesis is performed in the absence of phosphate (Table 5). 

 These algae were lightly centrifuged from their growth mediiom and 

 resuspended in water. If the algae were resuspended in .001 M 

 phosphate, no C^'^-labeled glycolate was formed in 40 seconds and 

 a 1 to 2 minute period was required before any glycolate-C^'* was 

 synthesized (G. Orth and N. E. Tolbert, unpublished). Whitting- 

 ham's data on glycolate excretion confirms this phenomenon (29). 

 However, the presence of phosphate in the medium does not prevent 

 the algae from ultimately synthesizing and excreting large amounts 



