Biosynthesis of porphyrins and related macrocycles. Part 20. Purification of deaminase and studies on its mode of action

A procedure is described for isolation of essentially pure deaminase from Euglena gracilis. Experiments with this enzyme prove (a) that it assembles four intact units of porphobilinogen; (b) the intermediate mono-, di-, tri-, and tetra-pyrroles are covalently bound to deaminase before release as the hydroxymethylbilane; (c) this bilane is built starting with ring-A and ending with ring-D and each of the four units of porphobilinogen is built into the tetrapyrrole at the same rate; (d) a very reactive tetrapyrrolic intermediate is formed (even more reactive than the hydroxymethylbilane) and evidence is given from trapping experiments that this reactive species is the linear tetrapyrrolic azafulvene (6).