Structure of porphobilinogen synthase from Pseudomonas aeruginosa in complex with 5-fluorolevulinic acid suggests a double Schiff base mechanism.
All natural tetrapyrroles, including hemes, chlorophylls and vitamin B12, share porphobilinogen (PBG) as a common precursor. Porphobilinogen synthase (PBGS) synthesizes PBG through the asymmetric condensation of two molecules of aminolevulinic acid (ALA). Crystal structures of PBGS from various sources confirm the presence of two distinct binding sites for each ALA molecule, termed A and P. We have solved the structure of the active-site variant D139N of the Mg2+-dependent PBGS from Pseudomonas aeruginosa in complex with the inhibitor 5-fluorolevulinic acid at high resolution. Uniquely, full occupancy of both substrate binding sites each by a single substrate-like molecule was observed. Both inhibitor molecules are covalently bound to two conserved, active-site lysine residues, Lys205 and Lys260, through Schiff bases. The active site now also contains a monovalent cation that may critically enhance enzymatic activity. Based on these structural data, we postulate a catalytic mechanism for P. aeruginosa PBGS initiated by a C-C bond formation between A and P-side ALA, followed by the formation of the intersubstrate Schiff base yielding the product PBG.
Frère
Frederic
F
Institute of Microbiology, Technical University Braunschweig, Spielmannstrasse 7, D-38106 Braunschweig, Germany.
Schubert
Wolf-Dieter
WD
Stauffer
Frédéric
F
Frankenberg
Nicole
N
Neier
Reinhard
R
Jahn
Dieter
D
Heinz
Dirk W
DW
D000622:Aminolevulinic Acid / Q000737:chemistry*; D001483:Base Sequence; D001665:Binding Sites; D002384:Catalysis; D002412:Cations; D018360:Crystallography, X-Ray; D007982:Levulinic Acids / Q000737:chemistry* / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D008274:Magnesium / Q000494:pharmacology; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D000623:Porphobilinogen Synthase / Q000737:chemistry*; D011485:Protein Binding; D011550:Pseudomonas aeruginosa / Q000201:enzymology*
C462259:5-fluorolevulinic acid; D002412:Cations; D007982:Levulinic Acids; D000622:Aminolevulinic Acid; D000623:Porphobilinogen Synthase; D008274:Magnesium; D008239:Lysine