bioRxiv Science⌕ Search

Biology subjects

Hosogi, N.

Publications and source records attributed to Hosogi, N..

2 recordsLinked to original sources

A representative of a ubiquitous bacterial lineage parasitically feeds on host RNA

Cellular metabolism is widely understood as an integrated network of redox reactions, energy conservation, and biosynthetic pathways. Here we show that across diverse prokaryotic lineages, loss of redox-associated functions is coupled with loss of nucleotide biosynthesis, raising a fundamental question of how such metabolically reduced organisms sustain cell growth. One of the most widespread and diverse lineages of prokaryotes--Minisyncoccota or Patescibacteriota--constitutes the majority of lineages exhibiting this pattern. To investigate how such organisms persist, we cultivated and characterized a representative of this lineage from a deep aquifer. The organism attaches to and penetrates growing bacterial host cells, and directly uptakes host RNA, concomitant with its depletion in the host. The metabolically reduced parasite cleaves host-derived RNA to directly supply cellular energy currencies and precursors for RNA/DNA synthesis, NTPs, without invoking canonical metabolic pathways. Codon usage in the parasite is complementary to that of its host, potentially minimizing translation of host-derived mRNA. Comparative genomics and phylogenetics indicate that these features are widespread and likely ancestral across the lineage. Exploitation of host RNA as a metabolic resource reveals a previously unrecognized metabolic strategy that demonstrates cellular metabolism can be sustained through direct utilization of informational macromolecules and thereby taps into an omnipresent energy reservoir, potentially supporting the environmental ubiquity of the metabolically reduced lineage.

microbiology↗

Use of phase plate cryo-EM reveals conformation diversity of therapeutic IgG with 50 kDa Fab fragment resolved below 6 angstrom

While cryogenic electron microscopy (cryo-EM) is fruitfully used for harvesting high-resolution structures of sizable macromolecules, its application to small or flexible proteins composed of domains like immunoglobulin (IgG) remain challenging. Here, we applied single particle cryo-EM to Rituximab, a therapeutic IgG mediating cancer cell toxicity, to explore its solution conformations. We found Rituximab molecules exhibited aggregates in cryo-EM specimens contrary to its solution behavior, and utilized a non-ionic detergent to successfully disperse them as isolated particles amenable to single particle analysis. As the detergent adversely reduced the protein-to-solvent contrast, we employed phase plate contrast to mitigate the impaired protein visibility. Assisted by phase plate imaging, we obtained a canonical three-arm IgG structure with other structures displaying variable arm densities co-existing in solution, affirming high flexibility of arm-connecting linkers. Furthermore, we showed phase plate imaging enables reliable structure determination of Fab to sub-nanometer resolution from ab initio, yielding a characteristic two-lobe structure that could be unambiguously docked with crystal structure. Our findings revealed conformation diversity of IgG and demonstrated phase plate was viable for cryo-EM analysis of small proteins without symmetry. This work helps extend cryo-EM boundaries, providing a valuable imaging and structural analysis framework for macromolecules with similar challenging features.

biochemistry↗