Dodecin Is the Key Player in Flavin Homeostasis of Archaea

Abstract

Flavins are employed to transform physical input into biological output signals. In this function, flavins catalyze a variety of light-induced reactions and redox processes. However, nature also provides flavoproteins with the ability to uncouple the mediation of signals. Such proteins are the riboflavin-binding proteins (RfBPs) with their function to store riboflavin for fast delivery of FMN and FAD. Here we present in vitro and in vivo data showing that the recently discovered archaeal dodecin is an RfBP, and we reveal that riboflavin storage is not restricted to eukaryotes. However, the function of the prokaryotic RfBP dodecin seems to be adapted to the requirement of a monocellular organism. While in eukaryotes RfBPs are involved in trafficking riboflavin, and dodecin is responsible for the flavin homeostasis of the cell. Although only 68 amino acids in length, dodecin is of high functional versatility in neutralizing riboflavin to protect the cellular environment from uncontrolled flavin reactivity. Besides the predominant ultrafast quenching of excited states, dodecin prevents light-induced riboflavin reactivity by the selective degradation of riboflavin to lumichrome. Coordinated with the high affinity for lumichrome, the directed degradation reaction is neutral to the cellular environment and provides an alternative pathway for suppressing uncontrolled riboflavin reactivity. Intriguingly, the different structural and functional properties of a homologous bacterial dodecin suggest that dodecin has different roles in different kingdoms of life.Flavins are a major class of cofactors that are able to accept and donate electrons as well as to absorb visible light. Flavins consist of a nonconserved aliphatic moiety, covalently linked to a conserved aromatic isoalloxazine ring ( In all flavins, the catalytically active unit is the isoalloxazine substructure (2-4). To handle the reactivity of the isoalloxazine ring, FMN and FAD are tightly bound to flavoenzymes. Finely tuned binding restricts the reaction spectrum of FMN and FAD to discrete, beneficial reaction pathways, preventing harmful side reactions. Reported functions of flavoenzymes include transferring electrons from and to reactive centers (e.g. respiratory chain) as well as employing light for the induction of radical reactions (e.g. DNA-photolyase) or conformational changes (e.g. phototropin) (5-8). In the biosynthesis, the full catalytic power of flavins has already formed at the stage of the FMN and FAD precursor RF (see Fig. 1). As RF is not used as an enzymatically active compound, it is, different to FMN and FAD, not integrated into flavoenzymes. To avoid a negative impact of uncontrolled flavin reactivity by free RF, nature established sequestering devices termed riboflavin-binding proteins (RfBPs). These proteins have been shown to store and distribute RF in eukaryotes, ensuring a fast supply of FMN and FAD (9, 10).The small flavoprotein dodecin from the archaeon Halobacterium salinarum was reported to bind RF with high affinity and to extensiv...

Ultrafast Excited-state Deactivation of Flavins Bound to Dodecin

“…With one ligand per 68 residues, dodecin represents the flavoprotein with highest flavin load known to date. Dodecin has also been found in bacteria where it was characterized as a binder of FMN (2)(3)(4).…”

“…Structures revealed a dodecameric fold, providing six identical binding pockets, which each enables binding of two antiparallel arranged flavins between two tryptophan residues building an aromatic tetrade arrangement ( Fig. 1A) (1)(2)(3)(4)(5)(6)(7).…”

“…Although both share the incorporation of antiparallel stacked flavin dimers, the bacterial dodecin incorporates si-si-stacked pairs of FMN, whereas archaeal dodecin binds re-re-stacked pairs of riboflavin. An alignment of approximately 100 dodecins reflects the separation of bacterial and archaeal dodecin on a sequence level (2). The physiological function of the bacterial dodecins is still not known.…”

“…RfbPs are high affinity binders, and reduce the photochemical reactivity of the highly reactive riboflavins by efficiently quenching light-induced excited states. Dodecin has recently been identified as a RfbP in archaea, representing the first protein of this function in prokaryotes (1,2). With one ligand per 68 residues, dodecin represents the flavoprotein with highest flavin load known to date.…”

Crystal structure of calcium dodecin (Rv0379), from Mycobacterium tuberculosis with a unique calcium‐binding site

“…The overall three-dimensional architecture of the Ca dodecin oligomer is similar to the Halobacterium salinarum dodecin (hsDodecin), a riboflavinbinding protein involved in riboflavin homeostasis. [14][15][16][17] The three subunits bind one calcium ion at the interface [ Fig. 1(b)], and the association of four equivalent sets of three subunits and a calcium leads to dodecameric oligomerization [ Fig.…”