Discovery and characterization of a novel family of prokaryotic nanocompartments involved in sulfur metabolism

Abstract

Prokaryotic nanocompartments, also known as encapsulins, are a recently discovered proteinaceous organelle-like compartments in prokaryotes that compartmentalize cargo enzymes. While initial studies have begun to elucidate the structure and physiological roles of encapsulins, bioinformatic evidence suggests that a great diversity of encapsulin nanocompartments remains unexplored. Here, we describe a novel encapsulin in the freshwater cyanobacterium Synechococcus elongatus PCC 7942. This nanocompartment is upregulated upon sulfate starvation and encapsulates a cysteine desulfurase enzyme via an N-terminal targeting sequence. Using cryo-electron microscopy, we have determined the structure of the nanocompartment complex to 2.2 Å resolution. Lastly, biochemical characterization of the complex demonstrated that the activity of the cysteine desulfurase is enhanced upon encapsulation. Taken together, our discovery, structural analysis, and enzymatic characterization of this prokaryotic nanocompartment provide a foundation for future studies seeking to understand the physiological role of this encapsulin in various bacteria.

Pore dynamics and asymmetric cargo loading in an encapsulin nanocompartment

“…In the H. ochraceum encapsulin the five-fold pore has a negative charge on the exterior of the encapsulin shell and positive charge on the interior in both the open and closed conformations (Figures S9 and S10). The closed conformation is consistent with observations from the crystal structure of the T. maritima encapsulin ( 7) and high resolution cryo-EM structures of other encapsulins (9,24,25). However, this is the first time that an 'open' pore-conformation has been observed in an encapsulin protein.…”

Nanotechnological Applications Based on Bacterial Encapsulins

“…The encapsulins are a family of~30 kDa bacterial and archaeal proteins that selfassemble into icosahedral compartments which range in size from 20 to 42 nm. These nanocompartments are similar to virus capsids in the icosahedral architecture of the container as well as in the protomer fold, which is structurally similar to capsid proteins of the Hong Kong 97 (HK97)-like virus lineage that includes the Caudovirales bacteriophages [6,[29][30][31][32].…”

“…There has been a gradual improvement in the algorithms for sequence similarity database searches and a continuous increase in the size and diversity of sequenced genomes of bacteria and archaeal microorganisms; this is producing a progressively sharper picture of the true abundance and diversity of encapsulins in nature [6,29,31,35]. The most recent computational database search identified over 6000 encapsulin-like systems in 31 bacterial and four archaeal phyla, grouped in four different families based on particle structure [32].…”

“…The evolutionary origin of encapsulins remains elusive; this is due in part to the low sequence similarity between encapsulins and members of the HK97-like virus lineage, which implies very old origin, and in part to the few atomic structures available. Based on phylogenetic and structural analyses, encapsulins are proposed to have a viral origin, possibly via "domestication" events of prophage HK97-type capsid proteins [6,[29][30][31][32]. Eukaryotic cells show evidence of similar events involving the Ty3/gypsy superfamily of retrotransposons, ancient mobile elements that are widely distributed and often abundant in their genomes, considered ancestral to modern retroviruses [45].…”

“…At present, eight encapsulins have been resolved at near-atomic resolution by X-ray crystallography or three-dimensional cryo-electron microscopy (3D cryo-EM). These are the encapsulins from the archaeon P. furiosus (EncPf; PDB entry 2e0z) [29], and from the bacteria T. maritima (EncTm; PDB entry 3dkt) [6], Myxococcus xanthus (EncMx; PDB entry 4pt2) [30], Quasibacil-lus thermotolerans (EncQt; PDB entry 6nj8) [49], Synechococcus elongatus (EncSe; PDB entry 6 × 8 m) [31], Mycolicibacterium hassiacum (EncMh; PDB entry 6i9g) [50], Mycobacterium smegmatis (EncMs; PDB entry 7boj) [51], and Haliangium ochraceum (EncHo) [52]. All these encapsulins except EncSe are close relatives and are grouped in the classical family 1; EncSe belongs to family 2A and has distinctive structural and functional features.…”