ABSTRACT
Studies on riboflavin biosynthesis in Bacillus subtilis were prompted by the isolation of mutants excreting substantial amounts of the vitamin. Light and heavy riboflavin synthase have been purified to homogeneity from cell extracts of the flavinogenic B. subtilis mutant H 94. The heavy enzyme is purified by gel permeation chromatography followed by batch crystallization. The light enzyme is purified by cation-exchange chromatography using a pH gradient followed by chromatography on phenyl Sepharose and hydroxyapatite. The quaternary structure of heavy riboflavin synthase can be experimentally modulated in a surprisingly wide range. The 1-MDa enzyme complex of B. subtilis is the only protein with lumazine synthase activity which has been studied in detail to date. Ligand binding studies had shown earlier that one pyrimidine or lumazine ligand can be bound per β subunit. The icosahedral β subunit capsid of riboflavin synthase is rather densely packed with the exception of the channels along the fivefold axes.
