Novel assembly of the SF370 strain of the important human pathogen serotype M1.

The important human pathogen causes a wide range of diseases. Strain SF370 was the first fully sequenced strain of , providing essential insights into the molecular mechanisms of disease. We present an improved genome assembly of strain SF370 from ATCC, which corrects long-standing errors in its genome.

The energy metabolism of in different trophic conditions.

The "knallgas" bacterium is attracting interest due to its extremely versatile metabolism. can use hydrogen or formic acid as an energy source, fixes CO the Calvin-Benson-Bassham (CBB) cycle, and grows on organic acids and sugars. Its tripartite genome is notable for its size and duplications of key genes (CBB cycle, hydrogenases, and nitrate reductases).

A prokaryotic Argonaute protein recruits a helicase-nuclease to degrade invading plasmids.

DdmDE is a novel plasmid defense system that was discovered in the seventh pandemic Vibrio cholerae strain of the biotype O1 EI Tor. In this issue of Cell, Yang and coworkers reveal the mechanisms underlying the assembly and activation of the DdmDE defense system.

Reflections from Nobel laureates in chemistry.

Since the first award in 1901, the Nobel Prize has come to signify the pinnacle of scientific achievement. In this Voices piece in the August special issue of Cell Chemical Biology entitled "Bridging chemistry and biology," we ask Nobel laureates to reflect on the impact the prize had on them. We learn how it affected their life or work, their outlook on science, the lessons learned, and their advice for the next generation of scientists.

Dynamics of diversified A-to-I editing in Streptococcus pyogenes is governed by changes in mRNA stability.

Adenosine-to-inosine (A-to-I) RNA editing plays an important role in the post-transcriptional regulation of eukaryotic cell physiology. However, our understanding of the occurrence, function and regulation of A-to-I editing in bacteria remains limited. Bacterial mRNA editing is catalysed by the deaminase TadA, which was originally described to modify a single tRNA in Escherichia coli.

Bacterial motility depends on a critical flagellum length and energy-optimised assembly.

Unlabelled: The flagellum is the most complex macromolecular structure known in bacteria and comprised of around two dozen distinct proteins. The main building block of the long, external flagellar filament, flagellin, is secreted through the flagellar type-III secretion system at a remarkable rate of several tens of thousands amino acids per second, significantly surpassing the rates achieved by other pore-based protein secretion systems. The evolutionary implications and potential benefits of this high secretion rate for flagellum assembly and function, however, have remained elusive.

Expanding the genetic toolbox for the obligate human pathogen .

Genetic tools form the basis for the study of molecular mechanisms. Despite many recent advances in the field of genetic engineering in bacteria, genetic toolsets remain scarce for non-model organisms, such as the obligatory human pathogen To overcome this limitation and enable the straightforward investigation of gene functions in , we have developed a comprehensive genetic toolset. By adapting and combining different tools previously applied in other Gram-positive bacteria, we have created new replicative and integrative plasmids for gene expression and genetic manipulation, constitutive and inducible promoters as well as fluorescence reporters for .

A versatile regulatory toolkit of arabinose-inducible artificial transcription factors for Enterobacteriaceae.

The Gram-negative bacteria Salmonella enterica and Escherichia coli are important model organisms, powerful prokaryotic expression platforms for biotechnological applications, and pathogenic strains constitute major public health threats. To facilitate new approaches for research and biotechnological applications, we here develop a set of arabinose-inducible artificial transcription factors (ATFs) using CRISPR/dCas9 and Arabidopsis-derived DNA-binding proteins to control gene expression in E. coli and Salmonella over a wide inducer concentration range.

Systems-Wide Site-Specific Analysis of Glycoproteins.

Glycosylation is one of the most common and complex post-translation modifications that influence the structural and functional properties of proteins. Glycoproteins are highly heterogeneous and exhibit site- and protein-specific expression differences. Mass spectrometry in combination with liquid chromatography has emerged as the most powerful tool for the comprehensive characterization of glycosylation.

Methodologies for bacterial ribonuclease characterization using RNA-seq.

Bacteria adjust gene expression at the post-transcriptional level through an intricate network of small regulatory RNAs and RNA-binding proteins, including ribonucleases (RNases). RNases play an essential role in RNA metabolism, regulating RNA stability, decay, and activation. These enzymes exhibit species-specific effects on gene expression, bacterial physiology, and different strategies of target recognition.

Automated Phosphopeptide Enrichment for Gram-Positive Bacteria.

Protein phosphorylation in prokaryotes has gained more attention in recent years as several studies linked it to regulatory and signaling functions, indicating importance similar to protein phosphorylation in eukaryotes. Studies on bacterial phosphorylation have so far been conducted using manual or HPLC-supported phosphopeptide enrichment, whereas automation of phosphopeptide enrichment has been established in eukaryotes, allowing for high-throughput sampling. To facilitate the prospect of studying bacterial phosphorylation on a systems level, we here established an automated Ser/Thr/Tyr phosphopeptide enrichment workflow on the Agilent AssayMap platform.

Time-Resolved Proteome Analysis of during Infection Reveals the Role of the AAA+ Chaperone ClpC for Host Cell Adaptation.

The cellular proteome comprises all proteins expressed at a given time and defines an organism's phenotype under specific growth conditions. The proteome is shaped and remodeled by both protein synthesis and protein degradation. Here, we developed a new method which combines metabolic and chemical isobaric peptide labeling to simultaneously determine the time-resolved protein decay and synthesis in an intracellular human pathogen.

Control of membrane barrier during bacterial type-III protein secretion.

Type-III secretion systems (T3SSs) of the bacterial flagellum and the evolutionarily related injectisome are capable of translocating proteins with a remarkable speed of several thousand amino acids per second. Here, we investigate how T3SSs are able to transport proteins at such a high rate while preventing the leakage of small molecules. Our mutational and evolutionary analyses demonstrate that an ensemble of conserved methionine residues at the cytoplasmic side of the T3SS channel create a deformable gasket (M-gasket) around fast-moving substrates undergoing export.

Survival Strategies of in Response to Phage Infection.

Bacteriophages exert strong evolutionary pressure on their microbial hosts. In their lytic lifecycle, complete bacterial subpopulations are utilized as hosts for bacteriophage replication. However, during their lysogenic lifecycle, bacteriophages can integrate into the host chromosome and alter the host's genomic make-up, possibly resulting in evolutionary important adjustments.

Distinct Polysaccharide Utilization Determines Interspecies Competition between Intestinal Prevotella spp.

Prevotella spp. are a dominant bacterial genus within the human gut. Multiple Prevotella spp.

Bridge helix arginines play a critical role in Cas9 sensitivity to mismatches.

The RNA-programmable DNA-endonuclease Cas9 is widely used for genome engineering, where a high degree of specificity is required. To investigate which features of Cas9 determine the sensitivity to mismatches along the target DNA, we performed in vitro biochemical assays and bacterial survival assays in Escherichia coli. We demonstrate that arginines in the Cas9 bridge helix influence guide RNA, and target DNA binding and cleavage.

Characterization of a transcriptional TPP riboswitch in the human pathogen .

Increasing evidence has demonstrated that regulatory RNA elements such as riboswitches (RS) play a pivotal role in the fine-tuning of bacterial gene expression. In this study, we investigated and characterized a novel transcriptional thiamine pyrophosphate (TPP) RS in the obligate human pathogen MC58 (serogroup B). This RS is located in the 5´ untranslated region upstream of gene, encoding a protein involved in TPP biosynthesis, an essential cofactor for all living beings.

Evolutionary classification of CRISPR-Cas systems: a burst of class 2 and derived variants.

The number and diversity of known CRISPR-Cas systems have substantially increased in recent years. Here, we provide an updated evolutionary classification of CRISPR-Cas systems and cas genes, with an emphasis on the major developments that have occurred since the publication of the latest classification, in 2015. The new classification includes 2 classes, 6 types and 33 subtypes, compared with 5 types and 16 subtypes in 2015.

Catalytically Active Cas9 Mediates Transcriptional Interference to Facilitate Bacterial Virulence.

In addition to defense against foreign DNA, the CRISPR-Cas9 system of Francisella novicida represses expression of an endogenous immunostimulatory lipoprotein. We investigated the specificity and molecular mechanism of this regulation, demonstrating that Cas9 controls a highly specific regulon of four genes that must be repressed for bacterial virulence. Regulation occurs through a protospacer adjacent motif (PAM)-dependent interaction of Cas9 with its endogenous DNA targets, dependent on a non-canonical small RNA (scaRNA) and tracrRNA.

CRISPR-Cas in Streptococcus pyogenes.

The discovery and characterization of the prokaryotic CRISPR-Cas immune system has led to a revolution in genome editing and engineering technologies. Despite the fact that most applications emerged after the discovery of the type II-A CRISPR-Cas9 system of Streptococcus pyogenes, its biological importance in this organism has received little attention. Here, we provide a comprehensive overview of the current knowledge about CRISPR-Cas systems from S.