Purification and Properties of l-Lysine Monooxygenase from Pseudomonas fluorescens
Marcia I. S. Flashner 1 and Vincent Massey 1
From the
1 From the Department of Biological Chemistry, The University of Michigan, Ann Arbor, Michigan 48104
l-Lysine monooxygenase has been isolated in nearly pure form from a lysine adapted strain of Pseudomonas fluorescens. The enzyme, which catalyzes the oxidative decarboxylation of l-lysine to form 
-aminovaleramide, has a molecular weight of approximately 246,000, and contains four subunits of equal size and four FAD prosthetic groups. Enzymic activity is greatly inhibited by a variety of buffers at low lysine concentrations, while at saturating l-lysine levels, this sensitivity is lost, and buffers have very little effect on the activity. The absorption spectrum is typical of a simple flavoprotein, with absorption maxima at 276, 385, and 462 nm. Two electron equivalents per mole are required to completely reduce the enzyme-bound flavin by dithionite, indicating that FAD is the only electron acceptor associated with the enzyme. During the course of reduction by dithionite, the semiquinone form of the flavin is produced. The ionization state of this species is a function of pH, with red or anionic semiquinone seen at pH values above 8 and the blue or neutral form at pH 5.5. l-Lysine monooxygenase, then, is one of the few flavoproteins exhibiting a pK near neutrality for the semiquinone oxidation state. The fluorescence properties of this enzyme are also unusual in that while the oxidized state is practically devoid of visible fluorescence, the reduced state exhibits flavin fluorescence, with an emission maximum at 505 nm and an excitation maximum at 360 nm. This reduced enzyme fluorescence is quenched by the addition of the substrates l-lysine or l-ornithine.
Submitted on August 20, 1973
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