Utilization and metabolism of urea during spore germination by Geotrichum candidum

Urea can serve as a sole N₂ source for spore germination of Geotrichum candidum which lakcs urease. However, when endogenous urea concentration exceeded 50 μmole/g dry wt., a significant inhibition of protein synthesis and spore germination occurred. At external urea concentration of 5 × 10^-3 M, approximately 95% of the urea-¹⁴C assimilated by the spores evolved as ¹⁴CO₂. Urea-¹⁴C incorporation showed temperature and pH dependence and obeyed saturation kinetics. The uptake of urea was completely inhibited by 10^-3 M NaN₃. A sharp increase in the accumulation of endogenous urea was detected when extracellular urea exceeded 6 × 10^-2 M. The latter increase was proportional to external substrate concentration until it reached about 1·5 × 10^-1 M. Most of the labelled C incorporated into germinated spores during 15 min feeding period with either urea-¹⁴C or NaH¹⁴CO₃ (with ammonia), was located in the cationic fraction. However, whereas citrulline, aspartate and glutamate were concurrently labelled immediately following NaH¹⁴CO₃ incorporation, citrulline was preferentially labelled when the germinated spores were fed with urea-¹⁴C. ATP:urea amidolyase (UALase) was purified approximately 126 fold from germinated spores of G. candidum. The enzymes had an optimum pH of 7·8 and the Km values for ATP and urea were 1-3 × 10^-3 M and 1·4 × 10^-4 M respectively. Avidin and KF were potent inhibitors of UALase activity. Attempts to demonstrate either enzyme induction by urea or enzyme repression by ammonia gave negative results.