Main physicochemical features of monofunctional flavokinase from Bacillus subtilis.
The main properties of a monofunctional riboflavin kinase from B. subtilis have been studied for the first time; the enzyme is responsible for a key reaction in flavin biosynthesis--the ATP-dependent phosphorylation of riboflavin with production of flavin mononucleotide. The active form of the enzyme is a monomer with molecular weight of about 26 kD with a strict specificity for reduced riboflavin. To display its maximum activity, the enzyme needs ATP and Mg(2+). During the phosphorylation of riboflavin, Mg(2+) could be partially replaced by ions of other bivalent metals, the efficiencies of which decreased in the series Mg(2+) > Mn(2+) > Zn(2+), whereas Co(2+) and Ca2+ had inhibiting effects. The flavokinase activity was maximal at pH 8.5 and 52 degrees C. ATP could be partially replaced by other triphosphates, their donor activity decreasing in the series: ATP > dATP > CTP > UTP. The Michaelis constants for riboflavin and ATP were 0.15 and 112 micro M, respectively. As compared to riboflavin, a tenfold excess of its analog 7,8-dimethyl-10-(O-methylacetoxime)-isoalloxazine decreased the enzyme activity by 30%. Other analogs of riboflavin failed to markedly affect the enzyme activity.
Solovieva
I M
IM
Department of Molecular and Radiation Biophysics, Konstantinov St. Petersburg Institute of Nuclear Physics, Russian Academy of Sciences, Gatchina, Leningrad Region, 188350, Russia. solovieva@em.uni-frankfurt.de
Tarasov
K V
KV
Perumov
D A
DA
D000255:Adenosine Triphosphate / Q000494:pharmacology; D001412:Bacillus subtilis / Q000201:enzymology*; D002413:Cations, Divalent / Q000494:pharmacology; D005486:Flavin Mononucleotide / Q000096:biosynthesis; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D012256:Riboflavin / Q000031:analogs & derivatives / Q000378:metabolism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D013696:Temperature
D002413:Cations, Divalent; D005486:Flavin Mononucleotide; D000255:Adenosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C022532:riboflavin kinase; D012256:Riboflavin