Sequence Determinants of Quaternary Structure in Lumazine Synthase

MARÍA SILVINA FORNASARI; DIEGO A. LAPLAGNE; NICOLÁS FRANKEL; ANA A. CAUERHFF; FERNANDO A. GOLDBAUM; JULIÁN ECHAVE

MOLECULAR BIOLOGY AND EVOLUTION

OXFORD UNIV PRESS

Año: 2004 vol. 21 p. 97 - 107

0737-4038

Riboflavin, an essential cofactor for all organisms, is biosynthesized in plants, fungi and microorganisms. Thepenultimate step in the pathway is catalyzed by the enzyme lumazine synthase. One of the most distinctive characteristicsof this enzyme is that it is found in different species in two different quaternary structures, pentameric and icosahedral,built from practically the same structural monomeric unit. In fact, the icosahedral structure is best described as a capsid oftwelve pentamers. Despite this noticeable difference, the active sites are virtually identical in all structurally studiedmembers. Furthermore, the main regions involved in the catalysis are located at the interface between adjacent subunitsin the pentamer. Thus, the two quaternary forms of the enzyme must meet similar structural requirements to achieve theirfunction, but, at the same time, they should differ in the sequence traits responsible for the different quaternary structuresobserved.Here, we present a combined analysis that includes sequence-structure and evolutionary studies to find the sequencedeterminants of the different quaternary assemblies of this enzyme. A data set containing 86 sequences of the lumazinesynthase family was recovered by sequence similarity searches. Seven of them had resolved three-dimensional structures.A subsequent phylogenetic reconstruction by maximum parsimony (MP) allowed division of the total set into twoclusters in accord with their quaternary structure. The comparison between the patterns of three-dimensional contactsderived from the known three-dimensional structures and variation in sequence conservation revealed a significant shiftin structural constraints of certain positions. Also, to explore the changes in functional constraints between the twogroups, site-specific evolutionary rate shifts were analyzed.We found that the positions involved in icosahedral contacts suffer a larger increase in constraints than the rest. Wefound eight sequence sites that would be the most important icosahedral sequence determinants. We discuss our resultsand compare them with previous work. These findings should contribute to refinement of the current structural data, tothe design of assays that explore the role of these positions, to the structural characterization of new sequences, and toinitiation of a study of the underlying evolutionary mechanisms.