Purification and characterization of flavokinase from Neurospora crassa.

The ATP-dependent phosphorylation of riboflavin to FMN by flavokinase is the key step in flavin biosynthesis. Flavokinase has been purified from a fungal source for the first time. The enzyme purified from a cell wall lacking mutant of Neurospora crassa, slime, is a monomer of M(r) 35.5 kDa with maximal activity at alkaline pH and high temperature (55 degrees C). The K(m) for both substrates is the lowest reported for flavokinase from any source so far (120 nM for riboflavin and 210 nM for MgATP2-). The enzyme exhibits preference for Mg2+ over Zn2+ as the essential activator and is also significantly activated by several cations. Activation by orthophosphate may be physiologically relevant for the intracellular regulation of flavokinase.

Activation of enzymatic catalysis.

Modulation of enzyme activity by inhibition and activation plays an important physiological role in regulation of cellular metabolism. Compared to the wealth of information available regarding inhibition of metabolic pathways, little is known about activation. Limited proteolysis of zymogens exemplifies irreversible activation. Reversible activation may involve post-translational modifications or dissociable binding of small molecules. Sometimes, chemical modification may also activate enzymes. The influence of small molecules on the reversible binding and activation of enzymes is summarized.