title: Pseudomonas aeruginosa contains a novel type V porphobilinogen synthase with no required catalytic metal ions.
abstract: Porphobilinogen synthases (PBGS) are metalloenzymes that catalyze the first common step in tetrapyrrole biosynthesis. The PBGS enzymes have previously been categorized into four types (I-IV) by the number of Zn(2+) and/or Mg(2+) utilized at three different metal binding sites termed A, B, and C. In this study Pseudomonas aeruginosa PBGS is found to bind only four Mg(2+) per octamer as determined by atomic absorption spectroscopy, in the presence or absence of substrate/product. This is the lowest number of bound metal ions yet found for PBGS where other enzymes bind 8-16 divalent ions. These four Mg(2+) allosterically stimulate a metal ion independent catalytic activity, in a fashion dependent upon both pH and K(+). The allosteric Mg(2+) of PBGS is located in metal binding site C, which is outside the active site. No evidence is found for metal binding to the potential high-affinity active site metal binding sites A and/or B. P. aeruginosa PBGS was investigated using Mn(2+) as an EPR probe for Mg(2+), and the active site was investigated using [3,5-(13)C]porphobilinogen as an NMR probe. The magnetic resonance data exclude the direct involvement of Mg(2+) in substrate binding and product formation. The combined data suggest that P. aeruginosa PBGS represents a new type V enzyme. Type V PBGS has the remarkable ability to synthesize porphobilinogen in a metal ion independent fashion. The total metal ion stoichiometry of only 4 per octamer suggests half-sites reactivity.
authors.affiliation: Institut für Organische Chemie und Biochemie, Albert-Ludwigs-Universität, Albertstrasse 21, 79104 Freiburg im Breisgau, Germany.
authors.forename: N
authors.initials: N
authors.lastname: Frankenberg
authors.forename: D
authors.initials: D
authors.lastname: Jahn
authors.forename: E K
authors.initials: EK
authors.lastname: Jaffe
mesh_terms: D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D001665:Binding Sites; D002247:Carbon Isotopes; D002384:Catalysis; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism; D008345:Manganese / Q000737:chemistry / Q000378:metabolism; D008670:Metals / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D000623:Porphobilinogen Synthase / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D013054:Spectrophotometry, Atomic
chemical_list: D002247:Carbon Isotopes; D008670:Metals; D008345:Manganese; D000623:Porphobilinogen Synthase; D008274:Magnesium