56 NITROGEN METABOLISM 



A series of well-controlled experiments has failed to find 

 any evidence for the participation of NHgOH in the fixation 

 of N 2 by Azotohacter and CI. pasteurianum [31, 33]. Hydrox- 

 ylamine in concentrations greater than about 2 /<g./ml. was 

 toxic, and growth, when it occurred, could be accounted for 

 in terms of NH3 produced by the spontaneous decomposi- 

 tion of NHgOH. Even with Ni^HgOH the results were diffi- 

 cult to interpret because the amount of nitrogen involved 

 was too small to permit significant determinations. The 

 oximes of a-ketoglutarate, oxaloacetate and pyruvate were 

 slowly utilized by CI. pasteurianum but not by Azoto- 

 hacter [34]. 



Pathways of N^-fixation 



There has been little support in recent years for the 

 fixation of nitrogen being explained in terms of an oxidative 

 pathway involving compounds such as nitrous oxide and 

 hyponitrous acid. There is now a considerable amount of 

 evidence that it is probably a process involving a number of 

 reductive mechanisms which terminate in the formation of 

 NH3 . Whilst the steps between molecular Ng and NH3 are 

 completely unknown some hypothetical pathways are shown 

 in Fig. 4.2 (details of other schemes will be found in 

 reference 44). Although the fixation of Ng by cell-free 

 systems has yet to be confirmed, past failures [24] may be 

 attributed, at least in part, to the harsh procedures employed 

 in the preparation of the extracts. 



In spite of their diverse nature, bacteria, blue-green algae 

 and leguminous plants all apparently fix nitrogen by 

 mechanisms which, if not identical, have at least many 

 features in common [43]. They all respond in a similar 

 manner to changes in the partial pressure of Ng, Hg and 

 CO and their K^ values for Ng are of the same order. In 

 view of the repeated failures to demonstrate Ng-fixation by 

 free-living cultures of Rhizobium, interest with regard to the 

 symbiotic system is centred around the question why 

 association with the leguminous plants should endow one 

 or other of the symbionts with this ability. A noticeable 

 feature of the association is the presence of a red haemo- 



