No abstract

CO2 is converted into biomass almost solely by the enzyme rubisco. The poor carboxylation properties of plant rubiscos have led to efforts that made it the most kinetically characterized enzyme, yet these studies focused on < 5% of its natural diversity. Here, we searched for fast‐carboxylating variants by systematically mining genomic and metagenomic data. Approximately 33,000 unique rubisco sequences were identified and clustered into ≈ 1,000 similarity groups. We then synthesized, purified, and biochemically tested the carboxylation rates of 143 representatives, spanning all clusters of form‐II and form‐II/III rubiscos. Most variants (> 100) were active in vitro, with the fastest having a turnover number of 22 ± 1 s−1—sixfold faster than the median plant rubisco and nearly twofold faster than the fastest measured rubisco to date. Unlike rubiscos from plants and cyanobacteria, the fastest variants discovered here are homodimers and exhibit a much simpler folding and activation kinetics. Our pipeline can be utilized to explore the kinetic space of other enzymes of interest, allowing us to get a better view of the biosynthetic potential of the biosphere.

Bacterial anti-phage defense systems are frequently clustered in microbial genomes, forming defense islands. This genomic property enabled the recent discovery of multiple defense systems based on their genomic co-localization with known systems, but the full arsenal of anti-phage mechanisms in bacteria is still unknown. In this study we report the discovery of 21 new defense systems that protect bacteria from phages, based on computational genomic analyses and phage infection experiments. We find multiple systems with protein domains known to be involved in eukaryotic anti-viral immunity, including ISG15-like proteins, dynamin-like proteins, and SEFIR domains, and show that these domains participate in bacterial defense against phages. Additional systems include protein domains predicted to manipulate DNA and RNA molecules, as well as multiple toxin-antitoxin systems shown here to function in anti-phage defense. The systems we discovered are widely distributed in bacterial and archaeal genomes, and in some bacteria form a considerable fraction of the immune arsenal. Our data substantially expand the known inventory of defense systems utilized by bacteria to counteract phage infection.

It has become clear in recent years that anti-phage defense systems cluster non-randomly within bacterial genomes in so-called “defense islands”. Despite serving as a valuable tool for the discovery of novel defense systems, the nature and distribution of defense islands themselves remain poorly understood. In this study, we comprehensively mapped the defense system repertoire of >1,300 strains of Escherichia coli, the most widely studied organism for phage-bacteria interactions. We found that defense systems are usually carried on mobile genetic elements including prophages, integrative conjugative elements and transposons, which preferentially integrate at several dozens of dedicated hotspots in the E. coli genome. Each mobile genetic element type has a preferred integration position but can carry a diverse variety of defensive cargo. On average, an E. coli genome has 4.7 hotspots occupied by defense system-containing mobile elements, with some strains possessing up to eight defensively occupied hotspots. Defense systems frequently co-localize with other systems on the same mobile genetic element, in agreement with the observed defense island phenomenon. Our data show that the overwhelming majority of the E. coli pan-immune system is carried on mobile genetic elements, explaining why the immune repertoire varies substantially between different strains of the same species.

In recent years it has become clear that anti-phage defence systems cluster non-randomly within bacterial genomes in so-called "defence islands". Despite serving as a valuable tool for the discovery of novel defence systems, the nature and distribution of defence islands themselves remain poorly understood. In this study, we comprehensively mapped the repertoire of defence islands within >1,300 strains of Escherichia coli, the most widely studied organism in terms of phage-bacteria interactions. We found that defence islands preferentially integrate at several dozens of dedicated integration hotspots in the E. coli genome. Defence islands are usually carried on mobile genetic elements including prophages, integrative conjugative elements and transposons, as well as on other genetic elements whose nature of mobilisation is unclear. Each type of mobile genetic element has a preferred integration position but can carry a diverse variety of defensive cargo. On average, an E. coli genome has 4.5 genomic hotspots occupied by a defence system-containing mobile element, with some strains possessing up to eight defensively occupied hotspots. Our data show that the overwhelming majority of the E. coli pan-immune system is carried on mobile genetic elements that integrate at a discrete set of genomic hotspots, and explains why the immune repertoire substantially varies between different strains of the same species.