Elucidation of a novel primitive pathway of aneorobic porphyrin biosynthesis

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Sulfate-reducing bacteria are broadly found in soil of ricefield, marine sediments, wastewater biofilms, animal guts, and the bacteria play important roles in the respective environments. Desulfovibrio species synthesize heme anaerobically by an unusual pathway including methylation of C-2 and C-7 positions of porphyrin ring. This research project aims to elucidate the whole pathway of heme biosynthesis and shed new light on evolution of porphyrin biosynthesis. The following results were obtained.(1) A new intermediate of (12, 18-didecarboxy)precorrin-2 was identified. Based on the structure, a tentative pathway of primitive heme biosynthesis was proposed.(2) To verify the proposed pathway in terms of enzymatic reactions, sensitive and rapid methods were developed to detect and quantify compounds possibly involved in the pathway such as glutamate-1-semialdehyde, sirohydrochlorin, (12,18-didecarboxy)sirohydrochlorin, and (2/7-monodecarboxymethyl)(12,18-didecarboxy)sirohydrochlorin.(3) To carry out strictly anaerobic enzyme reactions (OィイD22ィエD2 concentration less than 0.1μM), catechol 2,3-dioxygenase and its substrate catechol were used to measure and control OィイD22ィエD2 concentration in reaction mixtures. X-ray crystal structure of the enzyme in complex with acetone, a competitive inhibitor, was determined.(4) Cloning of the genes encoding enzymes responsible for heme biosynthesis has been tried systematically. Some of them will be soon elucidated.