Fig. 1.

Growth dependence on availability of heme and quantification of heme b in P. serpens and related flagellates. (A) Growth rate of P. serpens is the same without heme or when heme is supplied up to a concentration of 5 μM; 25 μM heme inhibits growth, likely attributable to toxic effects of free heme. Quite opposite dependence of growth on heme concentration is observed in the closely related C. fasciculata, which grows best when supplied with 25 μM heme and stops growing when heme concentration in the media is lowered to 1 μM. (B) Heme b extracted from equal numbers of cells from various kinetoplastids was separated by HPLC and detected by diode array detector. C. fasciculata was used as a related organism that possesses a complete set of respiratory complexes. It was grown in the same medium and supplemented with the same amount of heme (5 μM) as P. serpens. The bloodstream stage of T. brucei, which does not express its respiratory complexes III and IV in this life cycle stage, and thus functionally resembles P. serpens, was used as another control. The absence of respiratory complexes that normally consume most of heme is reflected in the much lower amount of extracted heme compared with C. fasciculata. The heme content in P. serpens is even lower than in T. brucei, which is in accordance with the lowest number of heme proteins found in the Phytomonas spp. genomes among all kinetoplastids (Table 1). Not even a trace amount of heme is detected in P. serpens grown without heme (dotted line). +H, with heme; −H, without heme.