No abstract

The penultimate step in the pathway of riboflavin biosynthesis is catalyzed by the enzyme lumazine synthase (LS). One of the most distinctive characteristics of this enzyme is the structural quaternary divergence found in different species. The protein exists as pentameric and icosahedral forms, built from practically the same structural monomeric unit. The pentameric structure is formed by five 18-kDa monomers, each extensively contacting neighboring monomers. The icosahedrical structure consists of 60 LS monomers arranged as 12 pentamers giving rise to a capsid exhibiting icosahedral 532 symmetry. In all lumazine synthases studied, the topologically equivalent active sites are located at the interfaces between adjacent subunits in the pentameric modules. The Brucella sp. lumazine synthase (BLS) sequence clearly diverges from pentameric and icosahedric enzymes. This unusual divergence prompted us to further investigate its quaternary arrangement. In the present work, we demonstrate by means of solution light scattering and x-ray structural analyses that BLS assembles as a very stable dimer of pentamers, representing a third category of quaternary assembly for lumazine synthases. We also describe by spectroscopic studies the thermodynamic stability of this oligomeric protein and postulate a mechanism for dissociation/unfolding of this macromolecular assembly. The higher molecular order of BLS increases its stability 20°C compared with pentameric lumazine synthases. The decameric arrangement described in this work highlights the importance of quaternary interactions in the stabilization of proteins.Riboflavin, an essential cofactor for all organisms, is biosynthesized in plants, fungi, and microorganisms. The penultimate step in the pathway is catalyzed by the enzyme lumazine synthase (LS).1 One of the most distinctive characteristics of this enzyme is the structural quaternary divergence found in different species. The protein exists as pentameric and icosahedral forms, built from practically the same structural monomeric unit. The structure of the monomer consists of four repeated ␤-strand/␣-helix motifs producing a sandwich of four parallel ␤-strands surrounded by four ␣-helices, two on each face of the ␤ sheet (1). In all LS studied, the topologically equivalent active sites are located at the interfaces between adjacent subunits in the pentameric modules (2).Bacilliaceae express a bifunctional enzyme complex with lumazine synthase and riboflavin synthase activity. Three ␣-subunits (riboflavin synthase) enclosed by 60 ␤-subunits (lumazine synthase) form a protein particle of ϳ1 MDa (1, 3). The three-dimensional structure of the lumazine synthase/riboflavin synthase from Bacillus subtilis complexed with a substrate analogue has been determined (4). This structure consists of 60 ␤-subunits (lumazine synthase monomers) arranged as 12 pentamers giving rise to a capsid exhibiting icosahedral 532 symmetry. Two other icosahedric LS have been described by x-ray crystallography. Spinach and thermophilic bacterial Aqui...