A virus capsid‐like nanocompartment that stores iron and protects bacteria from oxidative stress

Abstract

Living cells compartmentalize materials and enzymatic reactions to increase metabolic efficiency. While eukaryotes use membrane-bound organelles, bacteria and archaea rely primarily on protein-bound nanocompartments. Encapsulins constitute a class of nanocompartments widespread in bacteria and archaea whose functions have hitherto been unclear. Here, we characterize the encapsulin nanocompartment from Myxococcus xanthus, which consists of a shell protein (EncA, 32.5 kDa) and three internal proteins (EncB, 17 kDa; EncC, 13 kDa; EncD, 11 kDa). Using cryo-electron microscopy, we determined that EncA self-assembles into an icosahedral shell 32 nm in diameter (26 nm internal diameter), built from 180 subunits with the fold first observed in bacteriophage HK97 capsid. The internal proteins, of which EncB and EncC have ferritin-like domains, attach to its inner surface. Native nanocompartments have dense iron-rich cores. Functionally, they resemble ferritins, cage-like iron storage proteins, but with a massively greater capacity (~30,000 iron atoms versus ~3,000 in ferritin). Physiological data reveal that few nanocompartments are assembled during vegetative growth, but they increase fivefold upon starvation, protecting cells from oxidative stress through iron sequestration.

Prokaryotic nanocompartments form synthetic organelles in a eukaryote

“…Self-assembly of cargo-loaded encapsulin compartments in yeast Expression and self-assembly of encapsulin compartments in yeast was achieved using a plasmid containing the encapsulin gene EncA from M. xanthus 14 under control of the inducible GAL1 promoter (Fig. 1B).…”

“…Catalytic turnover of an encapsulated yeast enzyme Finally, enzyme-loaded encapsulins were shown to be viable nanoreactors for catalytic processes. The assembled encapsulin structure 14 contains small 5-10 Å sized pores, which in principle, can allow small molecule substrates and products to diffuse in and out of the compartment. The candidate enzyme chosen for encapsulation was Aro10p, a tetrameric pyruvate decarboxylase enzyme that is endogenous to yeast, participating in the catabolism of aromatic amino acids such as tyrosine (Fig.…”

“…In vivo protein encapsulation is guided by short targeting peptides (TPs), which are located at the C-termini of cargo proteins. A large variety of native cargo proteins has been identified in bacteria and archaea, including peroxidases and ferritin-like proteins involved in stress response pathways 11,[14][15][16] . Using Escherichia coli as a host, it has been shown that packaging of non-native proteins into the encapsulins from Thermotoga maritima and Brevibacterium linens can be achieved by fusion of targeting peptides to the intended cargo 17,18 .…”

“…Here, we present the construction of synthetic organelles in the yeast Saccharomyces cerevisiae, based on the Myxococcus xanthus encapsulin 14 . We show that encapsulin compartments can stabilize heterologous cargo proteins against degradation, co-localize proteins within their interior, and act as nanoreactors for housing enzymatic catalysts (Fig.…”

Labelling Bacterial Nanocages with Photo‐switchable Fluorophores

“…They are composed of 60 identical proteins that self‐assemble into cages, with a diameter ranging between 20 and 24 nm (Figure 1). Their structure has been compared to T =1 viral protein cages in terms of dimension and symmetry 3, 5. However, while viruses encapsulate genetic material, encapsulins enclose smaller proteins in their inner cavity, for instance a dye‐decolorizing peroxidase in B. linens encapsulins and a ferritin‐like protein in T. maritima encapsulins 1, 2.…”