The biosynthesis of uroporphyrinogen III: mechanism of action of porphobilinogen deaminase.
The biosynthesis of the uroporphyrinogen III macrocycle from porphobilinogen requires the sequential participation of two enzymes--porphobilinogen deaminase (1-hydroxymethylbilane synthase, EC 4.3.1.8) and uroporphyrinogen III synthase (cosynthase, EC 4.2.1.75). The product of the deaminase-catalysed reaction is a highly unstable 1-hydroxymethylbilane called preuroporphyrinogen which acts as the substrate for the uroporphyrinogen III synthase, resulting in the exclusive formation of uroporphyrinogen III. In the absence of the synthase, preuroporphyrinogen cyclizes spontaneously to give uroporphyrinogen I. Porphobilinogen deaminase contains a dipyrromethane cofactor that acts as a primer onto which the tetrapyrrole chain is built. The assembly process occurs in stages through enzyme-intermediate complexes, ES, ES2, ES3 and ES4. The negatively charged carboxylates of the cofactor, substrate and intermediate complexes interact with positively charged amino acid side chains in the catalytic cleft. Mutagenesis of conserved arginines has dramatic effects on the assembly of the dipyrromethane cofactor and on the tetrapolymerization process. During the polymerization, the enzyme changes conformation to accommodate the elongating pyrrole chain. The structure of the deaminase from Escherichia coli has been determined by X-ray crystallography at 1.9A resolution and gives important insight into the enzymic mechanism. Aspartate 84 plays a key role in catalysis and its substitution by glutamate reduces kcat by two orders of magnitude.
School of Biological Sciences, Queen Mary and Westfield College, University of London, UK.
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Jordan
D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D011163:Hydroxymethylbilane Synthase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016415:Sequence Alignment; D014576:Uroporphyrinogen III Synthetase / Q000378:metabolism*; D014577:Uroporphyrinogens / Q000096:biosynthesis*
D014577:Uroporphyrinogens; D011163:Hydroxymethylbilane Synthase; D014576:Uroporphyrinogen III Synthetase