Zinc and cadmium 5-aminolevulinate dehydratase. Metal-dependent pH profiles.
Native 5-aminolevulinic acid dehydratase contains zinc ions, which are essential for the enzymatic activity. Replacement of zinc by cadmium yielded an active enzyme whose kinetic parameters (kkat and Km) are similar to those of the zinc enzyme in the neutral pH range. However, the pH profiles of kcat and Km were different due to different pKa values. Two groups both with pKa values of 6.5 in the free zinc enzyme, but with pKa values of 7.0 in the cadmium enzyme were calculated from plots of log (kcat/Km) versus pH. On the other hand, the enzyme-substrate complex is controlled by one acidic group (zinc pKa = 6.0, cadmium pKa = 6.4) and one basis group (zinc pKa = 8.2, cadmium pKa = 7.7) as calculated from plots of log kcat versus pH. The Arrhenius plots for kcat of the two enzymes show no significant difference, the free energies of activation are 77.1 kJ/mol for the zinc and 76.8 kJ/mol for the cadmium enzyme. From this and from previous work it is concluded that the metal ions are located near the active site and influence the ionisations of essential amino acid residues. From the pH profiles of the modifying reaction and inhibition by diethylpyrocarbonate a histidinyl residue is inferred as one of the ionisable groups of the active site.
Fachbereich Biologie/Chemie, Universität Bremen.
M
M
Schlösser
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D
Beyersmann
D002104:Cadmium / Q000378:metabolism*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D000623:Porphobilinogen Synthase / Q000378:metabolism*; D013696:Temperature; D013816:Thermodynamics; D015032:Zinc / Q000378:metabolism*
D002104:Cadmium; D000623:Porphobilinogen Synthase; D015032:Zinc