Cloning of a Human cDNA for Protoporphyrinogen Oxidase by Complementation in Vivo of a hemG Mutant of Escherichia coli

• DOI: 10.1074/jbc.270.14.8076
• Publication Date: 2002-07-26T14:58:38Z
• Authors: Koichi Nishimura; Shigeru Taketani; Hachiro Inokuchi
• Keywords: Protoporphyrinogen oxidase

Protoporphyrinogen oxidase (PPO; EC 1.3.3.4) is the enzyme that catalyzes in penultimate step heme biosynthetic pathway. Hemes are essential components of redox enzymes, such as cytochromes. Thus, a hemG mutant strain Escherichia coli deficient PPO defective aerobic respiration and grows poorly even rich medium. By complementation with human placental cDNA library, we were able to isolate clone enhanced poor growth strain. The encoded gene for PPO. Sequence analysis revealed consists 477 amino acids calculated molecular mass 50.8 kilodaltons. deduced protein exhibited high degree homology over its entire length acid sequence by hemY Bacillus subtilis. NH2-terminal contains conserved forms dinucleotide-binding site many flavin-containing proteins. Northern blot synthesis 1.8-kilobase pair mRNA A homogenate monkey kidney COS-1 cells had been transfected much higher activity than an extract control cells, this was inhibited acifluorfen, specific inhibitor Furthermore, expressed vitro 51-kilodalton protein, after incubation isolated mitochondria found be located mitochondria, having just same size before, indication mitochondrial has no apparent transport-specific leader sequence.