Porphobilinogen deaminase [PBGD; porphobilinogen ammonia-lyase (polymerizing), EC 4.3

We have cloned and sequenced a cDNA clone coding for human erythrocyte porphobilinogen deaminase. It encompasses the translated region, part of the 5' and the 3' untranslated regions. The deduced 3tt amino acid sequence is consistent with the molecular weight and the partial amino-acid sequence of the NH2 terminal of the purified erythrocyte enzyme. Southern analysis of human genomic DNA shows that its gene is present as a single copy in the human genome and Northern analysis demonstrates the presence of a single size species of mRNA in erythroid and non-erythroid tissues and in several cultured cell lines. Quantitative determinations indicate that the amount of PBG-D mRNA is modulated both by the erythroid nature of the tissue and by cell proliferation, probably at the transcriptional level. INTRODUCTIONPorphobilinogen deaminase (PBG-D, EC t-3-1.8) is the third enzyme of the biosynthetic pathway leading to the production of heme. It catalyses the head to tail condensation of four molecules of the raonopyrrole porphobilinogen, to form the linear tetrapyrrole, hydroxymethylbilane which is then converted by uroporphyrlnogen III synthase to uroporphyrinogen III (1). In humans, deficiency in its activity is responsible for a dominant hereditary disease: Acute Intermittent Porphyria (A.I.P.) (2). Studies using antibodies specifically directed against human erythrocyte PBG-D show that inactive cross reacting immunological material is present in about 20$ of patients with this disorder but absent from the remainder (3). Therefore Acute Intermittent Porphyria appears to be, at the molecular level, a heterogeneous disorder.

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Although the erythroid-specific promoter of human porphobilinogen deaminase [PBGD] gene has no TATA box, transcription is initiated at a single nucleotide. Using 5' and 3' deletions and point mutations, we have identified an element, located around the initiation site, which is necessary and sufficient for 'in vitro' accurate initiation of transcription. This 15 bp element extends 1 bp 5' and 14 bp 3' from the initiation site. It is composed of two regions, a proximal region centred on the cap site and a distal region which bears homology with the TdT initiator element. We show that a nuclear factor, present both in erythroid and non erythroid cells, binds the distal PBGD initiator element. Lack of heat inactivation suggests that initiation of transcription mediated by this element is not TFIID dependent. By transfection into erythroid cells, we also show that the proximal PBGD initiator element is essential for the selection of the initiation site but not for the regulation of transcription of the PBGD erythroid promoter during erythroid differentiation.